After 65 years, research is still fun.

Annu. Rev. Anim. Biosci. 2013.1:1-20. Downloaded from www.annualreviews.org by University of Chicago Libraries on 09/28/13. For personal use only.

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After 65 Years, Research Is Still Fun William Hansel Liberty Hyde Bailey Emeritus Professor, Cornell University, Ithaca, New York 14853 Gordon Cain Professor Emeritus, Louisiana State University, Baton Rouge, Louisiana 70803 Professor, Pennington Biomedical Research Center, Louisiana State University System, Baton Rouge, Louisiana 70808; email: [email protected]

Annu. Rev. Anim. Biosci. 2013. 1:1–20

Keywords

First published online as a Review in Advance on December 13, 2012

corpus luteum, pituitary, hypothalamus, cancer, progesterone

The Annual Review of Animal Biosciences is online at animal.annualreviews.org

Abstract

This article’s doi: 10.1146/annurev-animal-031412-103722 Copyright © 2013 by Annual Reviews. All rights reserved

In 1946, at the end of World War II, I entered graduate school at Cornell University, where I remained for 44 years. During that time, my laboratory produced more than 300 publications in the field of reproductive biology, including studies on nutrition and reproduction, the role of the hypothalamus in pituitary gonadotropin release, corpus luteum formation and function, hormone assays, and estrous cycle synchronization. At age seventy, I retired from Cornell and accepted the Gordon Cain Endowed Professorship at Louisiana State University, where I continued my work on the bovine corpus luteum and added research on the collection, maturation, in vitro fertilization, and culture of bovine oocytes. In 1994, I moved to the Pennington Biomedical Research Center and soon thereafter started the research that led to development of the lytic peptide–gonadotropin conjugates, which target and destroy cancer cell membranes. I am continuing my work on the development of targeted cancer cell drugs and, yes, research is still fun!

1

INTRODUCTION


In May 1967, Carl Hartman presented a famous after-dinner speech at the Third Brook Lodge Workshop at Augusta, Michigan, entitled “Research Should Spell Fun.” Forty-five years later, after spending a total of 66 years in research and teaching, I can confirm that research is indeed fun. I entered Cornell University as a graduate assistant in Animal Physiology in February 1946. At that time, I was still using crutches as a result of injuries suffered during World War II. I spent five years in the Army during the war, most of it as an infantry company commander in General Patton’s US Third Army. I was wounded in the fighting through the heavily fortified Saar-Moselle River Triangle, the southern anchor of Germany’s Siegfried Line. At Cornell, I was fortunate to have an exceptionally gifted team of mentors, who influenced greatly my development as a scientist (Figure 1). My graduate committee chairman, S.A. Asdell, was one of the three leading domestic animal reproduction physiologists in the nation. He received his PhD degree at Cambridge University under the tutelage of F.H.A. Marshall, author of the first textbook of physiology. Asdell later received the coveted Marshall Medal for his research in reproductive physiology. Asdell insisted that my thesis research be completely original; thus, he offered no advice as to what the next experiment should be but always made every effort to secure the animals and equipment for whatever experiments I designed. Years later, I realized that this “sink or swim” philosophy of graduate student training contributed greatly to my ability to analyze a research problem and design appropriate experiments to solve it.

Sydney A. Asdell

H. H. Dukes

Kenneth L. Turk

C. M. McCay

Figure 1 My mentors: Sydney A. Asdell, H.H. Dukes, Kenneth L. Turk, and C.M. McCay.

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The second member of my graduate committee, H.H. Dukes, Chairman of the Physiology Department in the Veterinary College, was a great teacher. He was the author of the most widely used physiology text, Physiology of the Domestic Animals. The demonstrations of physiological principles devised by Dr. Dukes are unmatched, even to this day. In later years, I was privileged to contribute two chapters to his book. The third member of my graduate committee, K.L. Turk, Chairman of the Animal Science Department, was well known to me, because he served as my undergraduate advisor at the University of Maryland before World War II. Dr. Turk grew up in Missouri and was a strong believer in applying research results to practical problems in agriculture. I remember him best for his oftenasked question, “William, what have you done for the farmers of New York State this week?” I still look for applications of my research to agriculture or medicine. A fourth person at Cornell who greatly influenced my career was C.M. McCay, the discoverer of the principle that growth retardation, produced by restriction of caloric intake, prolongs the life span of rats. This principle was later established in nearly all species in which it was tested and is now widely regarded as one of the most important biological experiments ever conducted! McCay and his students occupied the laboratory adjacent to Dr. Asdell’s, and I often talked to him, enrolled in his courses, and, at times, assisted in some of his experiments. McCay was clearly a man ahead of his time, and I learned from him how to isolate and solve difficult research problems. A fifth person who influenced my development at Cornell was J.B. Sumner, a Nobel Prize winner for his work in isolating the first enzyme, urease, from jack beans. Sumner had lost one arm, but he carried out routine laboratory procedures with remarkable dexterity. He taught a great course in biochemistry, which ended with a final exam consisting of one question: “Write all you know about proteins!” This was not an easy task, even in those days when much less was known. These mentors, along with the high-quality graduate students they attracted, made Cornell an ideal training ground for postwar students in physiology and nutrition.

NUTRITION AND REPRODUCTION Not surprisingly, a major effort was made at Cornell in the early post–World War II period to extend the results of McCay’s studies (1, 2) on the effects of growth retardation on life span in rats to domestic animals. In 1935, Asdell & Crowell (3) showed that the occurrence of first estrus and ovulation were delayed in rats fed calorie-restricted diets. In our first experiment (4, 5), we fed Holstein heifers from birth to 80 weeks of age on 60%, 100%, and 140% of recommended total digestible nutrient (TDN) allowances. Heifers fed on the high plane of nutrition first came into estrus at 37.4 6 7.1 weeks of age, compared with 49.1 6 6.3 weeks of age for heifers fed the normal diet and 72 weeks or more for heifers fed the low level of nutrition (Figure 2). Methods for measuring steroids and gonadotropins in blood or tissues were not available at that time, but we were able to measure pituitary growth hormone (GH) and pituitary thyroid stimulating hormone (TSH) by bioassays to show that levels of GH were high in young animals and decreased with increasing age and that TSH levels were higher in heifers fed on the high plane than on the low plane (5). In later studies, Reid et al. (6, 7) found that cattle fed TDN-restricted diets from birth to first calving lived longer and had higher lifetime milk production than cattle fed higher TDN allowances. These were the first experiments to show that McCay’s findings extended to a nonrodent species, and they had a significant influence on the feeding and breeding of dairy and beef cattle. Later, the ability of restricted caloric intake to increase longevity was demonstrated in many species including rhesus monkeys (8), Drosophila (9), yeast (10), and Caenorhabditis elegans (11). However, it soon became evident from the classic studies of Kennedy & Mitra (12) and Frisch & McArthur (13) that dietary fat and obesity also play direct roles in female reproduction. www.annualreviews.org

After 65 Years, Research Is Still Fun

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Age at first estrus (weeks) Figure 2 Average body weights and ages at first estrus of Holstein heifers fed low (60%), medium (100%), or high (140%) recommended total digestible nutrient allowances from birth to 80 weeks of age. The Ragsdale standard is shown for comparison. Sixty animals (20 trios) were randomly allotted to slaughter ages of 16, 32, 48, 64, and 80 weeks within the three treatment levels. An additional four calves were slaughtered at 0–1 weeks of age (4).

These studies also indicated the need for a minimal amount of fat for normal reproductive development. With the discovery of leptin, produced only by fat cells, it became evident that adipose tissue may act as an endocrine organ (14). Leptin, as well as insulin, exerts inhibitory control on neuropeptide Y–producing neurons in the arcuate nucleus of the hypothalamus (15). In recent studies, it has been shown that the epididymal fat pad may produce a locally acting factor responsible for maintaining spermatogenesis in hamsters (16).

THE ROLE OF THE HYPOTHALAMUS IN GONADOTROPIN RELEASE FROM THE PITUITARY Until 1950, the anterior pituitary was considered the sole conductor of the endocrine orchestra, whose instruments included follicle stimulating hormone (FSH), luteinizing hormone (LH), prolactin, TSH, adrenocortical stimulating hormone, and GH. However, in 1949, Everett, Sawyer & Markee, among others, produced strong evidence for the existence of a neurogenic (hypothalamic) factor in the preovulatory surge of LH secretion in the rat (17). Donovan (18) summarized the results of these early studies. In 1951, we reported blockage of ovulation in the cow by atropine injections (19), strong evidence for the existence of a neurogenic mechanism in LH release and ovulation. In a series of experiments (20–23), we established that the preovulatory surge of LH release from the bovine anterior pituitary occurs during the first six hours of estrus, even though ovulation does not occur until 10–12 h after the end of a 16–18 h estrus. Years later, these early findings were confirmed when accurate, repeatable radio-immunoassays for measuring LH in bovine plasma were developed (24). In 1953, we (Armstrong & Hansel) discovered that daily injections of oxytocin on days 2–6 of the bovine estrous cycle prevented normal corpus luteum (CL) formation and resulted in shortened estrous cycles approximately 10 days in length (25). When we presented these data at a meeting of the world’s leading reproductive biologists at West Point, New York, in July 1959, they caused 4

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a major debate between those who believed that the hypothalamus produces a LH-releasing hormone (LHRH) that is transported to the anterior pituitary by the portal vascular system and Sir Solly Zuckerman, chairman of the conference and longtime proponent of the belief that the hypothalamus has “nothing to do with LH release and ovulation” (26). We believed that oxytocin exerted its effect through the uterus, which proved to be the case (27), and, with help from Jack Everett, Alan Parkes, and others, we presented a convincing argument for the existence of a hypothalamic hormone that causes pituitary LH release and ovulation. This discussion marked a change in our concepts of control of pituitary gland function and resulted in more National Institutes of Health support of studies of the hypothalamus. In 1971, Schally (28) and others achieved the isolation and identification of a gonadotropin-releasing hormone (GnRH or LHRH) of hypothalamic origin, a discovery that led to the award of a Nobel Prize for Schally.

Early work with rodents indicated that prolactin is responsible for formation of the CL and the maintenance of progesterone secretion (29). In 1964, we reported that only LH, or LH-containing pituitary preparations, was capable of overcoming the lytic effects of concurrently administered oxytocin (day 2–6) on bovine CL weights as well as progesterone contents and concentrations (30). Inactivation of LH in these preparations by incubating them with 6M urea abolished these effects. Our conclusion that LH is the major luteotropic hormone was challenged by other researchers (31), who reported that injections of human chorionic gonadotropin failed to increase plasma progesterone concentrations or prolong the life span of the CL. This challenge prompted us to perform several additional experiments, which demonstrated clearly that LH is the major luteotropic hormone. We showed that single injections of LH on the sixteenth day of the estrous cycle prolonged the functional life of the CL and the length of the estrous cycle (32). We then demonstrated that LH-containing preparations and crude anterior pituitary extracts significantly increased CL weights and progesterone contents in hysterectomized heifers (27). Finally, in 1967, we showed that progesterone synthesis in vitro by bovine luteal tissue slices is stimulated by highly purified bovine LH (Figure 3). LH alone stimulated progesterone production to the same degree as a combination of all six anterior pituitary hormones (33).

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ROLE OF THE UTERUS IN CORPUS LUTEUM REGRESSION


The essentiality of the uterus for CL regression was demonstrated by early studies showing that hysterectomy resulted in prolonged periods of CL maintenance and continued secretion of progesterone (27, 34, 35). In a search to find the luteolytic substances produced by the uterus, we extracted bovine endometrial tissue and tested various fractions for luteolytic activity in pseudopregnant hamsters. This work led to the isolation of arachidonic acid (AA) (36, 37), the precursor of the prostaglandins. A role for prostaglandins in bovine CL regression was first suggested by Babcock in 1965, in the discussion following the presentation of our paper on luteotropic and luteolytic mechanisms in bovine corpora lutea at the Second Brook Lodge Workshop, sponsored by the Upjohn Company (38). After summarizing the effects of the prostaglandin family of compounds known at that time, he wondered whether prostaglandins from the uterus could have a luteolytic effect. During the same year, we obtained prostaglandin from the Upjohn Company and tested its ability to cause luteolysis when injected directly into the corpora lutea of normal and hysterectomized heifers. Unfortunately, the compound proved to be luteotropic rather than luteolytic (39). Later, we realized that we had injected PGE2 rather than PGF2a in this experiment, and in 1972, McCracken et al. (40) identified PGF2a as the uterine luteolysin in sheep. The idea that a uterine luteolysin reaches the ipsilateral ovary and CL by way of a countercurrent mechanism in the ovarian pedicle, in which the ovarian artery and the utero-ovarian vein are tightly adherent, arose in conversations with Dr. James Goding in 1967. These conversations concerned the significance of small vessels (vasa vasori) seen in longitudinal histological slides of the bovine ovarian pedicle. In 1970, Dr. Goding’s research team (41) reported at the Second Annual Meeting of the Australian Society for Reproductive Biology that separation of the ovarian artery and the utero-ovarian vein of the ewe, and insertion of a fold of peritoneum between the two, resulted in CL maintenance. In the same report, these researchers found that infusions of PGF2a into the ovarian arterial circulation of ewes with ovarian transplants caused plasma P to decline rapidly, which resulted in estrus within three days. In 1971, McCracken, Baird & Goding (42) reported that transplanting the CL-bearing ovary and grafting it into the neck, where the ovarian artery was anastomosed to the carotid and the ovarian vein to the jugular, resulted in prolonged CL maintenance in the ewe, and they identified PGF2a as the uterine luteolysin. We demonstrated the preferential transfer of PGF2a to the ovarian artery following its intrauterine administration in cattle (43). At this point, it seemed clear that PGF2a, produced in the uterus and transferred to the ovarian artery and thus the ipsilateral ovary and CL, is the mechanism for luteal regression in ruminants. Intramuscular injections of PGF2a in Holstein heifers on days 6–16 of the estrous cycle caused CL regression, a decline in plasma P concentrations, and a return to estrus; injections on days 2–5 were not effective (44). These early experiments led to the development of methods for estrous cycle synchronization, now widely used in dairy and beef cattle management, and to the widespread use of PGF2a to treat anestrus in dairy and beef cattle.

IN VITRO STUDIES OF BOVINE LUTEAL TISSUE AND CELLS In 1971, we summarized results of our in vitro studies with washed, minced luteal tissues at the Third Karolinska Symposium on In Vitro Methods in Reproductive Cell Biology (45). These studies confirmed that LH is the major luteotropic hormone. In 1981, Koos & Hansel (46) published a paper describing a method based on the use of unit gravity sedimentation for the separation of the small, theca interna–derived and the large, granulosa cell–derived steroidogenic cells in the bovine CL (Figure 4). This report and the 6

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FLA488 nm Figure 4 Separation of large granulosa cell–derived and small theca cell–derived luteal cells by fluorescence activated cell sorting (48). The endothelial cells of the corpus luteum vasculature are contained in the group of very small cells (< 10 mm).

subsequent report by Alila et al. (47), in which flow cytometry was used to further purify the cell preparations (Figure 4), allowed the cells to be studied individually and in combination and was used in hundreds of subsequent studies of steroidogenesis in luteal tissues. Dr. Koos received the Young Investigator Award of the Society for the Study of Reproduction for this work. Alila & Hansel (48) characterized the origin of the two cell types through the use of specific monoclonal antibodies. Dr. Alila also received the Young Investigator Award from the Society for the Study of Reproduction for his work. Studies that used the separated cell system soon revealed that, in contrast to its action in vivo, PGF2a in vitro is luteotropic, acting primarily on the small luteal cells (49). Other studies (50) showed that LH induces rapid increases in intracellular Caþþ that differ in magnitude and profile between small and large luteal cells and that progesterone production in luteal cells involves Caþþ/calmodulin/protein kinase C–dependent mechanisms (Figure 5). In the Hammond Memorial Lecture in 1985 (51), we developed the concept that progesterone synthesis in the small cells is controlled primarily by the cAMP system and that elevated intracellular calcium diminishes cAMP-mediated progesterone synthesis in these cells. In the James Goding Memorial Lecture in 1986 (52), I reported that the protein kinase C system exerts its effect on total cell preparations by stimulating the production of prostacyclin.

THE ROLE OF ARACHIDONIC ACID IN CORPUS LUTEUM FUNCTION In 1976, we infused AA for 24 h directly into a small branch of the ovarian artery of Holstein heifers, in which all connections between the ovarian pedicle and the ipsilateral uterine horn were severed. Plasma progesterone concentrations were markedly lowered during a 70-h period www.annualreviews.org


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LH (ng/ml) Annu. Rev. Anim. Biosci. 2013.1:1-20. Downloaded from www.annualreviews.org by University of Chicago Libraries on 09/28/13. For personal use only.

Figure 5 The effect of calcium deprivation on progesterone production by small theca-derived luteal cells in response to increasing concentrations of luteinizing hormone (LH) (50).

following infusion in the treated animals but were maintained in control vehicle–treated heifers. Because we had previously isolated AA, the precursor of PGF2a, from bovine endometrial tissue, these results suggested that the bovine CL can make its own PGF2a if supplied with sufficient AA (53). This proved to be the case; the ability of bovine luteal tissue to produce prostaglandins, both in vivo and in vitro, has been established fully (54–56). In 1992, Lafrance & Hansel (57) showed that AA elicited a significant release of oxytocin from the large cells and progesterone from both the large and small luteal cells. AA appears to play a key role in bovine CL function and needs further study, especially in light of two subsequent studies in which we were unable to demonstrate elevated concentrations of ovarian arterial PGF2a when concentrations of PGF2a in the utero-ovarian vein were elevated. In 1975, Shemesh & Hansel (58) measured levels of PGF in bovine endometrium as well as uterine venous and ovarian arterial and jugular plasma during the entire estrous cycle. PGF levels in the ovarian artery were not significantly higher than those of the peripheral blood, even at times when very high levels were found in the uterine vein. PGF levels in the ovarian artery did not rise significantly at any time during the cycle. In 1980, Milvae & Hansel (59) inserted uterine venous and ovarian arterial catheters into heifers on day 3 of the estrous cycle and treated them with oxytocin on days 4, 5, and 6. The oxytocin treatments depressed jugular vein progesterone concentrations by day 8 and increased uterine venous PGF concentrations between 30 and 240 min after each injection. However, no increases in PGF concentrations in ovarian arterial blood were found, even when uterine venous concentrations were highly elevated. These data do not support the concept of veno-arterial transfer of PGF in oxytocin-treated animals.

NITRIC OXIDE AND CORPUS LUTEUM REGRESSION These and other data suggested to us that a compound downstream from the AA metabolites might be involved in CL regression. Others had the same idea. In 1994, Pate (60) showed that apoptotic cell death is initiated by macrophages. Meidan and her coworkers (61) demonstrated that endothelin production by luteal blood vessels is stimulated by PGF2a. Endothelins, especially ET-1, synthesized and secreted by endothelial cells in the CL vasculature inhibit progesterone production by the luteal cells. 8

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Knowing that the bovine CL cells produce large amounts of nitric oxide (NO), we decided to test the hypothesis that NO is a downstream mediator of prostaglandin-induced CL regression. For these experiments, we used a microdialysis technique, first described by Jarry et al. (62), and installed as described by Blair et al. (63), to administer treatments and collect dialysate samples. Intraluteal perfusion of a NO synthase blocker, NLv-nitro-L-arginine methyl ester (L-NAME), on days 17 and 18 of the estrous cycle elevated progesterone levels in the perfusate and prolonged the length of the cycle through day 25 (64). These data indicate that NO plays an important role in the initiation of luteal regression in cattle. In subsequent studies, conducted in cooperation with scientists at the University of Warmia and Mazury Faculty of Veterinary Medicine and the Institute of Animal Reproduction and Food Research, Polish Academy of Science, Olsztyn, Poland, we showed that administration of a NO synthase inhibitor counteracts PGF2a-induced luteolysis (65, 66). These effects could not be demonstrated in pure cultures of either small or large luteal cells perfused with a NO donor or a NO synthase inhibitor, which suggests that endothelial cells may be required for the NO effect (67). Collectively, these data indicate clearly that NO is an important component of the luteolytic mechanism; it probably acts downstream from PGF2a and endothelin. The control of CL regression remains an active area of research, and controversies still exist. The key role of the CL in reproduction dictates that intensive research efforts in this area will continue. Furthermore, if we understood fully the mechanism(s) that bring about the reduction of a 5–6-g bovine CL that secretes large amounts of progesterone to less than 1 g of inactive tissue that is incapable of steroidogenesis, within a three-day period, we would likely be able to develop more effective anticancer drugs.

ESTROUS CYCLE SYNCHRONIZATION As artificial insemination techniques developed, there was an increasing demand for development of a technique for synchronizing the cycles of beef and dairy cattle and sheep, so that large numbers of animals could be inseminated on a single day. The idea that progesterone injections might be used to control the release of LH from the pituitary and thus the length of the estrous cycle traces to early studies reported in 1950 by Trimberger & Hansel (68) and Ulberg (69). These early experiments were not very successful; the variability in the time from last injection to the onset of estrus was very large, and conception rates of the cattle at the controlled estrus were low. Nevertheless, they established the principle that progesterone is the key to controlling cycle length. In 1959, at the West Point meeting of the world’s leaders in research in reproduction, referred to above, Gregory Pincus and his collaborators presented their data on the use of oral progestational compounds as contraceptives. We obtained several of these compounds and carried out many experiments in which these progestins were incorporated into the water supply or mixed in grain and fed to groups of dairy heifers or beef cattle. The first reports of successful synchronization of estrous cycles by orally effective progestational compounds appeared in 1960. Hansel & Malven (70) reported successful synchronization of 32 Hereford cows after feeding 500–968 mg of 6-methyl-17-acetoxy progesterone (MAP) per animal per day for 20 days. Nellor et al. (71) reported inhibition of estrus and ovulation after feeding MAP to beef heifers for 15–20 days. These early reports, which attracted worldwide attention, were followed by larger-scale trials involving 400 or more beef cattle. We reported results of these studies in 1966 at the Thirteenth Easter School of the University of Nottingham (72). MAP and CAP (6-chloro-D6-dehydro-17 acetoxy progesterone) were fed in pelleted feed to beef cattle. Of the MAP-fed beef cattle, 75% came into estrus by the third day following withdrawal of MAP, and 49% conceived when www.annualreviews.org


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inseminated with frozen semen, compared with 60% in the controls. CAP, a more potent progestin, also produced effective cycle synchronization, but it resulted in a lower conception rate (35%). At the same meeting, T.J. Robinson (73) reported successful cycle synchronization in ewes after withdrawal of progestin-bearing intravaginal sponges. Two progestins, SC9800 and MAP, were most effective in synchronizing the cycles and resulted in the highest conception rates. These pioneering studies stimulated great interest in cycle synchronization, and hundreds of research papers appeared in the literature in the next decade. In some cases, the sponges were replaced by progesterone-releasing intravaginal devices (PRIDs) designed for use in sheep and cattle, and in 1972 we reported at the Seventh International Congress on Animal Reproduction and Artificial Insemination in Munich, Germany (74), that administration of PGF2a can produce synchronization of estrous cycles in cattle. By 1978, we had developed an effective shortened method for cycle synchronization in cattle based on a single injection of PGF2a on the sixth day of a seven-day PRID treatment (PRID7-PGF2a6) (75). Finally, in 1984, we reported successful insemination of cycle-synchronized Holstein heifers at a preset time, without checking them for estrus (76). The seven-day period of PRID insertion remains an integral part of new methods being developed for fixed-time insemination of cyclesynchronized cattle. A major addition to the PRID-PGF2a method is the injection of GnRH at the time of PRID insertion and again at the time of insemination (77). Pregnancy rates for beef cattle inseminated by these methods at a fixed time after cycle regulation are nearly 70% and do not differ from pregnancy rates in control groups of cattle inseminated after normal estrous cycles.

STUDIES WITH BOVINE IN VITRO–FERTILIZED EMBRYOS In 1989, I served as Chairman of the National Organizing Committee for the Second Symposium on Genetic Engineering of Animals held at Cornell University and, with the able assistance of Barbara Weir, edited the proceedings (78). This remarkable experience added a new dimension to my research interests. When I retired from Cornell in 1990 and moved to Louisiana State University as the Gordon Cain Professor of Physiology, I gained access to the University’s wellequipped Embryo Laboratory, organized and administered by Dr. Robert Godke. This sequence of events resulted in a series of investigations over a five-year period (1993–1998) designed to improve the methods for harvesting, fertilizing, and culturing bovine oocytes. In our first study, we found that platelet-derived growth factor (PDGF) is required for development of the cocultured bovine embryo beyond the eight-cell stage (79). This so-called block stage was thought to represent the transition of genomic control from maternal to zygotic. In subsequent studies (80), we showed that the beneficial effects of culturing embryos in groups, rather than singly, were due to PDGF produced by the embryos in culture and that development of embryos beyond the 16-cell stage required transforming growth factor in the medium. These studies resulted in development of a chemically defined culture medium capable of supporting development of in vitro–fertilized bovine embryos cultured singly to the blastocyst stage (81, 82). Perhaps the most important of these studies of bovine embryo development was the discovery that addition of a NO scavenger, or an inhibitor of NO synthesis, significantly enhanced development to the blastocyst stage (83, 84). These studies resulted in the issue of two patents. In 1998, in cooperation with R.D. Randel at the Texas Agricultural Experiment Station, Overton, we found that high environmental temperature and humidity resulted in a marked decline in the quality of oocytes for Bos taurus cows and a marked decrease in their in vitro developmental capabilities. In contrast, oocytes from Bos indicus cows exhibited normal morphology and yielded a high percentage of blastocysts when cultured, regardless of the temperature (85). 10

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STUDIES WITH NONRUMINANT DOMESTIC ANIMALS Our work with the bovine CL was supported for many years by the National Institutes of Health, US Department of Agriculture (USDA) competitive grants, and the Binational Agricultural Research and Development (BARD) Fund as well as by several USDA-sponsored Regional Research Programs. Despite this strong impetus to work on bovine reproduction, we were able to expand our reproductive research activities to several other domestic animals, including the dog, mink, and alpaca. In the bitch, Concannon et al. (86) measured plasma levels of LH, estrogen, and progesterone throughout the entire ovarian cycle, as we had done previously in cattle (24). In the mink, we showed in 1980 that treatment with prolactin during the period of embryonic diapause advanced implantation and hastened the onset of luteal phase progesterone secretion (87), and in 1981 we established that rising levels of prolactin, induced by increasing hours of daylight, are solely responsible for ending the embryonic diapause (88). In the alpaca, an induced ovulator, we measured levels of plasma progesterone after mating and found that levels after infertile mating were not maintained, as had been assumed, but declined between 8 and 13 days (89). This rapid decline is the result of an active luteolytic process that involves the uterus (90). These fundamental findings were immediately useful for breeders of all three species; mink breeders found that a second mating during the embryonic diapause yields a 10–15% increase in the number of kits born. Alpaca breeders realized that animals that failed to conceive to first breeding could be rebred successfully as early as 13 days after the first breeding. Dog breeders used the hormone profile to design more effective methods to produce a fertile estrus and to treat infertility.

TARGETING CANCER CELLS BY REPRODUCTIVE HORMONES AND THEIR RECEPTORS In 1997, a series of events occurred that completely changed the direction of my research. During that year, my wife died of ovarian cancer, and I came to realize the inadequacy of the currently available anticancer drugs. During the same period, I attended a symposium in Olsztyn, Poland, at which C.V. Rao (91) reported LH and CG receptor expression by several cancer cell lines. At this point, I realized that compounds consisting of segments of the b chain of LH, CG, or FSH, coupled with an anticancer drug, might be used to target and destroy cancer cells that express the appropriate receptor. I also realized that the lytic peptides that Dr. Enright and I had tested previously as contraceptives for use in animals were the candidates of choice for the anticancer moiety of the molecule. On returning to the Pennington Biomedical Center, Dr. Enright and I, with the help of Dr. Carola Leuschner and Martha Juban, began to synthesize and test LH (CG), FSH, and LHRH– lytic peptide conjugates in the widely used nude mouse tumor model. The lytic peptides are defined as positively charged, linear, alpha-helical, amphipathic, membrane-disrupting compounds containing less than 40 amino acids (92). The synthetic lytic peptides that we selected (Hecate, Phor14, and Phor21) were each conjugated to segments of the b chain of CG or FSH or to LHRH. These conjugates (Hecate-bCG, Phor14-bCG, Phor21-bCG, LHRH-Hecate, and LHRH-Phor21) are described in greater detail by Hansel et al. (93). Phor14 and Phor21 contain two and three 7–amino acid sequences (KFAKFAK), respectively. Phor21 is more potent than Phor14. The lytic peptide conjugates are metabolized rapidly and are not antigenic. Because they act at the level of the cell membrane, they are not subject to the multiplex drug resistance phenomenon, in which cells develop the ability to excrete cytotoxic compounds by a transmembrane efflux pump (92). Because their activity is not dependent on destruction of all rapidly dividing cells, they lack many of the undesirable side effects of other www.annualreviews.org


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chemotherapeutic drugs. In 2001, we published our first papers (94, 95) on the targeted destruction of androgen-sensitive and -insensitive prostate cancer cells and xenografts through LH (CG) receptors. These papers were followed by a report in 2003 (96) in which we showed that LHRH-Hecate also targeted and destroyed human prostate cancer cells in vivo and in vitro. Similarly, we found in 2002 that a lytic peptide–bCG conjugate was effective in reducing tumor weights in mice bearing ovarian cancer xenografts (97). In 2003 and 2007, we reported that membrane-destroying lytic peptide conjugates were also capable of destroying breast cancer cells in vitro and in vivo (93, 98). Bodek et al. (99) reported in 2005 that a Hecate-bCG conjugate targeted Leydig cell tumors in transgenic mice. Recently, we found that a LHRH–lytic peptide conjugate is effective in reducing tumor weights and volumes in pancreatic cancer cell tumors (100). All of these cancer cells overexpress either CG or LHRH receptors, and all of the lytic peptide conjugates act primarily by destroying cell membranes and causing cell necrosis (101, 102). These conjugates have a remarkable ability to seek out and destroy metastatic cells in lymph nodes, bone, lungs, and other organs (93) (Figure 6). In a recent study (100), we showed that pretreatment of nude mice bearing pancreatic cancer cell xenografts with FSH enhances the ability of a LHRH-lytic peptide to cause tumor regression by upregulating LHRH receptors. Such methods have the potential to ensure that patients selected for treatment express adequate receptors. Esperance Pharmaceuticals Inc. has acquired rights to develop the lytic peptide conjugates as drugs to treat cancers. This company, with a recent infusion of funds from a large international pharmaceutical company, has completed phase I clinical trials and is currently in phase II clinical trials with its lead compound, EP-100, a LHRH–lytic peptide conjugate. At this point, we shifted our attention to development of drugs that improve apoptosis, because truly effective cancer treatments likely will be able to cause necrosis as well as optimize the apoptotic mechanisms. We found such a compound in curcumin, the active ingredient of the widely used Asian spice turmeric. Large doses of curcumin administered to mice by gavage were shown to

1,200 0

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*

*

*

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Phor21-βCG-ala (mg/kg) Lower spine 200

100

0

Control

0.2

2

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* P < 0.005 versus control

Figure 6 Luciferase-positive (metastatic) cells in homogenates of ribs and spines of nude mice bearing breast cancer cell (MDA-MB-435S.luc) tumors and treated with three dose levels of a lytic peptide conjugate. P < 0.005 versus control.

12

Hansel

ASSESSMENT OF FUTURE DEVELOPMENTS IN THE ANIMAL INDUSTRIES In 1954, in cooperation with Kenneth McEntee and Peter Olafson, pathologists in the Cornell Veterinary College, we isolated and identified highly chlorinated napthalenes as the causative agents of bovine hyperkeratosis (107, 108). Affected animals lacrimated continuously, developed thick skins resembling armor-plate, and were unable to convert carotene to vitamin A. The disease spread rapidly and, at first, was thought to be of viral origin. We isolated the causative agent, chlorinated napthalenes, from bread crumbs obtained from a bakery; it was also found in a wood preservative, in heat-resistant lubricants and fire-retardant paints. When the chlorinated napthalenes were removed from these products, the disease disappeared rapidly. I cite these experiments because they were among the first to isolate and solve a problem of contamination of our environment and food supply with a noxious agent. Problems of this nature are complex and increasing in frequency; their solution will require teamwork, and animal and veterinary scientists likely will play key roles. As the limitations of rodents, particularly mice, as models for use in studies of human cancer and obesity/diabetes become increasingly evident (109, 110), the need for better animal models becomes increasingly urgent. Use of the White Leghorn laying hen as a model for ovarian cancer

Tumor weight changes from baseline (g)


downregulate cancer cell proliferation and antiapoptotic gene products through suppression of the nuclear transcription factor NF-kb (103). Unfortunately, because of its poor solubility, humans cannot ingest enough curcumin to be effective. We conjugated curcumin with LHRH and found the conjugate to be highly soluble. When injected intravenously into nude mice bearing pancreatic cancer cell tumors, LHRH-curcumin caused a significant reduction in tumor weights and volumes (104) (Figure 7). LHRH-curcumin may become an effective drug for treatment of pancreatic ductal adenomas and other LHRHexpressing cancers, especially when used in combination with the LHRH–lytic peptide conjugates. Use of receptor-binding sequences of the b chains of the gonadotropins and LHRH, or its analogs, as targeting agents for cancer cells is increasing (105, 106), and we may expect further development of these promising new drugs.

1.4

a

b

LHRH

1.2 1.0

LHRH-curcumin

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0.6 0.4 0.2 0.0

** Untreated

LHRH

LHRHcurcumin

** Baseline

Treatment groups Figure 7 (a) Mean 6 standard-error tumor weight changes from baseline in nude mice bearing pancreatic cancer cell (MIA PaCa-2) xenografts, after treatment with luteinizing hormone–releasing hormone (LHRH) alone and with LHRH-curcumin conjugate, and in untreated controls. Baseline tumor weights were obtained from a group of tumor-bearing mice necropsied before the beginning of treatment. (b) Representative cancer cell tumors of nude mice treated with LHRH, LHRH-curcumin conjugate–treated mice, and untreated control mice ( P < 0.01) (104).



13

(111) is a good example of what can be accomplished in this field. Animal and veterinary scientists should contribute greatly to this emerging area of research. In 1986, in the William Henry Hatch Memorial Lecture (112), I predicted: “By the year 2000 we will have developed an animal breeding system in which artificial insemination has been replaced by a system of artificial inembryonation. In this system, highly trained technicians will place embryos into the uteri of animals whose estrous cycles have been regulated so that they are synchronized with the developmental stage of the embryo. The embryos transferred will have been produced by in vitro fertilization of ova from genetically superior females with sperm from genetically superior males. In most cases, the embryos will have been sexed, split and stored in a frozen state until used. Beginning about the year 2000, it was predicted that genetically engineered embryos containing genes that will result in faster growth rates, leaner carcasses, greater disease resistance


and improved lactational performances are likely to become available.”

In 1992, we reported that most of these goals had been achieved (113). However, in a special issue of Animal Reproduction Science entitled “Potential and Implications of Application of Reproductive Technologies,” it was necessary to point out that, although the technologies for achieving these goals had been developed, the animal industries had been slow to adopt them (114). This was largely because of the inefficiencies of the techniques involved, such as oocyte maturation, in vitro fertilization, embryo culture, and cloning. It suggested also that the best hope for increasing the impact of these technologies on the animal industries lies in developing efficient ways to mature, harvest, store, and fertilize in vitro the large numbers of primordial oocytes present in the ovaries of all farm animals. In other reports in the same issue, Paterson et al. (115) reported that cloning by the methods used to produce the famous sheep Dolly is inefficient—less than 4% of the embryos became viable offspring—and Seidel (116) reported the commercialization of the sexing of bovine sperm by flow cytometry. Hasler (117) pointed out that an improvement in the superovulation technique is needed for the embryo transfer industry to continue to grow. Since that time, progress in improving the efficiency of in vitro maturation has been slow, but a significant recent paper (118) reports a new method for oocyte in vitro maturation that markedly improves oocyte development. Since the birth of Dolly, reported by Wilmut et al. (119) in 1997, it has been known that the nuclear transfer of fully developed somatic cells (SCNT) could be used to produce live animals with the same genome as the somatic cell. The number of species in which SCNT has been achieved has increased greatly since 1997, but the efficiency of the procedure has not. Recent studies (120–22), in which mouse, human, and bovine fibroblasts have been reprogrammed to pluripotency by a cocktail of transcription factors (Oct 4, Sox2, cMyc, and Klf4), offer hope that induced pluripotent stem cells can be used in nuclear transfers.

DISCLOSURE STATEMENT The author serves as a member of the Scientific Advisory Boards for Esperance Pharmaceuticals, Inc. and K-94 Discoveries, Inc.

ACKNOWLEDGMENTS The work discussed was accomplished by more than one hundred pre- and postdoctoral students, many of whom are not cited because of limitations of space. I apologize for these omissions. The long-term assistance of my laboratory manager, Ray Saatman, is gratefully acknowledged. 14

Hansel

Financial support from Gordon Cain for the cancer research described is also gratefully acknowledged. The long-term administrative support of Cornell University, Louisiana State University, and the Pennington Biomedical Research Center is acknowledged, as are the numerous funding agencies that supported my research, including the National Institutes of Health, the US Department of Agriculture, the National Science Foundation, the Binational Agricultural Research and Development Fund, and Esperance Pharmaceuticals, Inc., among others.


LITERATURE CITED 1. McCay CM, Crowell MF. 1934. Prolong the life span. Sci. Mon. 39:405–14 2. McCay CM, Crowell MF, Maynard LA. 1935. The effect of retarded growth upon the length of the life span and upon the ultimate body size. J. Nutr. 10:63–79 3. Asdell SA, Crowell MF. 1935. The effect of retarded growth upon sexual development of rats. J. Nutr. 10:13–24 4. Sorenson AM, Hansel W, Hough WH, Armstrong DT, McEntee K, Bratton RW. 1959. Causes and prevention of reproductive failures in dairy cattle. I. The influence of underfeeding and overfeeding on growth and development of Holstein heifers. Cornell Univ. Agric. Exp. Stn. Bull. 936:3–37 5. Armstrong DT, Hansel W. 1956. The effect of age and plane of nutrition on growth hormone and thyrotropic hormone content of pituitary glands of Holstein heifers. J. Anim. Sci. 15:640–49 6. Reid JT, Loosli JK, Trimberger GW, Turk KL, Asdell SA, Smith SE. 1957. Progress report on a study of the effect of plane of nutrition upon reproductive and productive performance of Holstein cattle. J. Dairy Sci. 40:610–11 7. Reid JT, Loosli JK, Trimberger GW, Turk KL, Asdell SA, Smith SE. 1964. Causes and prevention of reproductive failures in dairy cattle. IV. The effect of plane of nutrition during early life on growth, reproduction, health, and longevity of Holstein cows. 1. Birth to fifth calving. Cornell Univ. Agric. Exp. Stn. Bull. 987:6–27 8. Colman RJ, Anderson RM, Johnson SC, Kastman EK, Kosmatka KJ, et al. 2009. Caloric restriction delays disease onset and mortality in rhesus monkeys. Science 325:201–4 9. Partridge L, Piper MD, Mair W. 2005. Dietary restriction in Drosophila. Mech. Ageing Dev. 126:938–50 10. Guarente L. 2005. Calorie restriction and SIR2 genes—towards a mechanism. Mech. Ageing Dev. 126: 923–28 11. Houthoofd K, Vanfleteren JR. 2006. The longevity effect of dietary restriction in Caenorhabditis elegans. Exp. Gerontol. 41:1026–32 12. Kennedy GC, Mitra J. 1963. Body weight and food intake as initiating factors for puberty in the rat. J. Physiol. 166:408–18 13. Frisch RE, McArthur JW. 1974. Menstrual cycles: fatness as a determinant of minimum weight for height necessary for their maintenance or onset. Science 185:949–51 14. Hansel W. 2010. The essentiality of the epididymal fat pad for spermatogenesis. Endocrinology 151:5565–67 15. Kalra SP, Xu B, Duke MG, Kalra PA. 1998. Nutritional infertility: mismanagement in central neuropeptidergic and peripheral insulin-leptin signaling. In Nutrition and Reproduction, ed. W Hansel, G Bray, DH Ryan, pp. 25–40. Baton Rouge: La. State Univ. Press 16. Chu Y, Huddleston GG, Clancy AN, Harris RB, Bartness TJ. 2010. Epididymal fat is necessary for spermatogenesis, but not testosterone production or copulatory behavior. Endocrinology 151:5669–79 17. Everett JW, Sawyer CH, Markee JE. 1949. A neurogenic timing factor in control of the ovulatory discharge of luteinizing hormone in the cyclic rat. Endocrinology 44:234–50 18. Donovan BT. 1967. Hypothalamic control of reproductive processes. In Control of Reproduction in the Female Mammal: Proceedings of the Thirteenth Easter School in Agricultural Science, ed. GE Lamming, EC Amoroso, pp. 3–29. London: Butterworths 19. Hansel W, Trimberger GW. 1951. Atropine blockage of ovulation in the cow and its possible significance. J. Anim. Sci. 10:719–25



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20. Jubb KV, McEntee K. 1955. Observations on the bovine pituitary gland. II. Architecture and cytology with special reference to basophil cell function. Cornell Vet. 45:593–641 21. Hansel W, Trimberger GW. 1952. The effect of progesterone on ovulation time in dairy heifers. J. Dairy Sci. 35:65–70 22. Hough WH, Bearden HJ, Hansel W. 1955. Further studies on factors affecting ovulation in the cow. J. Anim. Sci. 14:739–45 23. Hansel W. 1953. Neurogenic factors in ovulation. Iowa State Coll. J. Sci. 28:1–8 24. Hansel W, Echternkamp SE. 1972. Control of ovarian function in domestic animals. Am. Zool. 12:225–43 25. Armstrong DT, Hansel W. 1958. Alteration of the bovine estrous cycle with oxytocin. J. Dairy Sci. 42:533–42 26. Zuckerman S. 1963. Introduction. In Mechanisms Concerned with Conception, ed. CG Hartman, pp. 9– 18. New York: Pergamon Press 27. Malven PV, Hansel W. 1964. Ovarian function in dairy heifers following hysterectomy. J. Dairy Sci. 47:1388–93 28. Schally AV, Arimura A, Kastin AJ, Matsuo H, Baba Y, et al. 1971. Gonadotropin-releasing hormone: One polypeptide regulates secretion of luteinizing and follicle-stimulating hormones. Science 173:1036– 38 29. Malven PV, Hansel W, Sawyer CH. 1967. A mechanism antagonizing the luteotrophic action of exogenous prolactin in rats. J. Reprod. Fertil. 13:205–12 30. Simmons KR, Hansel W. 1964. Nature of the luteotropic hormone in the bovine. J. Anim. Sci. 23:136–41 31. Schomberg DW, Coudert SP, Short RV. 1967. Effects of bovine luteinizing hormone and human chorionic gonadotrophin on the bovine corpus luteum in vivo. J. Reprod. Fertil. 14:277–85 32. Donaldson LE, Hansel W. 1965. Prolongation of life span of the bovine corpus luteum by single injections of bovine luteinizing hormone. J. Dairy Sci. 48:903–4 33. Hansel W, Seifart KH. 1967. Maintenance of the luteal function in the cow. J. Dairy Sci. 50:1948–58 34. Wiltbank JN, Casida LE. 1956. Alterations of ovarian activity by hysterectomy. J. Anim. Sci. 15:134–40 35. Anderson LL, Neal FC, Malampy RM. 1961. Hysterectomy and ovarian function in beef heifers. Am. J. Vet. Res. 23:794–802 36. Lukaszewska JH, Hansel W. 1970. Extraction and partial purification of luteolytic activity from bovine endometrial tissue. Endocrinology 86:261–70 37. Hansel W, Shemesh M, Hixon J, Lukaszewska J. 1975. Extraction, isolation and identification of a luteolytic substance from bovine endometrium. Biol. Reprod. 13:30–37 38. Hansel W. 1966. Luteotropic and luteolytic mechanisms in bovine corpora lutea. J. Reprod. Fertil. S1:33–48 39. Brunner MA, Donaldson LE, Hansel W. 1969. Exogenous hormones and luteal function in hysterectomized and intact heifers. J. Dairy Sci. 52:1849–54 40. McCracken JA, Carson JC, Glow ME, Goding JR, Baird DT, et al. 1972. Prostaglandin F2a identified as a luteolytic hormone in sheep. Nat. New Biol. 238:129–34 41. Barrett S, Blockley MA, Brown JM, Cumming IA, Goding JR, et al. 1971. Initiation of the oestrous cycle in the ewe by infusions of PGF 2 alpha-to the autotransplanted ovary. J. Reprod. Fertil. 24:136–37 42. McCracken JA, Baird DT, Goding JR. 1971. Factors affecting the secretion of steroids from the transplanted ovary in the sheep. Recent Prog. Horm. Res. 27:537–82 43. Hixon JE, Hansel W. 1974. Evidence for preferential transfer of prostaglandin F2a to the ovarian artery following intrauterine administration in cattle. Biol. Reprod. 11:543–52 44. Beal WE, Milvae RA, Hansel W. 1980. Oestrous cycle length and plasma progesterone concentrations following administration of prostaglandin F-2a early in the bovine oestrous cycle. J. Reprod. Fertil. 59:393–96 45. Hansel W. 1971. Survival and gonadotrophin responsiveness of luteal cells in vitro. Acta Endocrinol. Suppl. 153:295–317 46. Koos R, Hansel W. 1981. The large and small cells of the bovine corpus luteum: ultrastructural and functional differences. In Dynamics of Ovarian Function, ed. NB Schwartz, M Heinzicker-Dunn, pp. 197–203. New York: Raven

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47. Alila HW, Dowd JP, Corradino RA, Harris WV, Hansel W. 1988. Control of progesterone production in small and large bovine luteal cells separated by flow cytometry. J. Reprod. Fertil. 82:645– 55 48. Alila HW, Hansel W. 1984. Origin of different cell types in the bovine corpus luteum as characterized by specific monoclonal antibodies. Biol. Reprod. 31:1015–25 49. Alila HW, Corradino RA, Hansel W. 1989. Differential effects of luteinizing hormone on intracellular free Ca2þ in small and large bovine luteal cells. Endocrinology 124:2314–20 50. Alila HW, Davis JS, Dowd JP, Corradino RA, Hansel W. 1990. Differential effects of calcium on progesterone production in small and large bovine luteal cells. J. Steroid Biochem. 36:687–93 51. Hansel W, Dowd JP. 1986. Hammond Memorial Lecture: new concepts of the control of corpus luteum function. J. Reprod. Fertil. 78:755–68 52. Hansel W, Alila HW, Dowd JP, Yang XZ. 1987. Control of steroidogenesis in small and large bovine luteal cells. Aust. J. Biol. Sci. 40:331–47 53. Hansel W, Fortune JE. 1978. The applications of ovulation control. In Control of Ovulation, ed. DB Crighton, NB Haynes, GR Foxcroft, GE Lamming, pp. 237–63. London: Butterworths 54. Milvae RA, Hansel W. 1983. Prostacyclin, prostaglandin F2a and progesterone production by bovine luteal cells during the estrous cycle. Biol. Reprod. 29:1063–68 55. Shemesh M, Hansel W. 1975. Arachidonic acid and bovine corpus luteum function. Exp. Biol. Med. 148:243–46 56. Shemesh M, Hansel W. 1975. Stimulation of prostaglandin synthesis in bovine ovarian tissues by arachidonic acid and luteinizing hormone. Biol. Reprod. 13:448–52 57. Lafrance M, Hansel W. 1992. Role of arachidonic acid and its metabolites in the regulation of progesterone and oxytocin release from the bovine corpus luteum. Exp. Biol. Med. 201:106–13 58. Shemesh M, Hansel W. 1975. Levels of prostaglandin F (PGF) in bovine endometrium, uterine venous, ovarian arterial and jugular plasma during the estrous cycle. Exp. Biol. Med. 148:123–26 59. Milvae RA, Hansel W. 1980. Concurrent uterine venous and ovarian arterial prostaglandin F concentrations in heifers treated with oxytocin. J. Reprod. Fertil. 60:7–15 60. Pate JL. 1994. Cellular components involved in luteolysis. J. Anim. Sci. 72:1884–90 61. Meidan R, Milvae RA, Weiss S, Levy N, Friedman A. 1999. Intraovarian regulation of luteolysis. J. Reprod. Fertil. Suppl. 54:217–28 62. Jarry H, Einspanier A, Kanngiesser L, Dietrich M, Pitzel L, et al. 1990. Release and effects of oxytocin on estradiol and progesterone secretion in porcine corpora lutea as measured by an in vivo microdialysis system. Endocrinology 126:2350–58 63. Blair RM, Saatman R, Liou SS, Fortune JE, Hansel W. 1997. Roles of leukotrienes in bovine corpus luteum regression: an in vivo microdialysis study. Exp. Biol. Med. 216:72–80 64. Jaroszewski JJ, Hansel W. 2000. Intraluteal administration of a nitric oxide synthase blocker stimulates progesterone and oxytocin secretion and prolongs the life span of the bovine corpus luteum. Exp. Biol. Med. 224:50–55 65. Jaroszewski JJ, Skarzynski DJ, Hansel W. 2003. Nitric oxide as a local mediator of prostaglandin F2ainduced regression in bovine corpus luteum: an in vivo study. Exp. Biol. Med. 228:1057–62 66. Skarzynski DJ, Jaroszewski JJ, Bah MM, Deptula KM, Barszczewska B, et al. 2003. Administration of a nitric oxide synthase inhibitor counteracts prostaglandin F2-induced luteolysis in cattle. Biol. Reprod. 68:1674–81 67. Jaroszewski JJ, Skarzynski DJ, Blair RM, Hansel W. 2003. Influence of nitric oxide on the secretory function of the bovine corpus luteum: dependence on cell composition and cell-to-cell communication. Exp. Biol. Med. 228:741–48 68. Trimberger GW, Hansel W. 1950. Conception rate and ovarian function following estrus control by progesterone injections in dairy cattle. J. Anim. Sci. 14:224–32 69. Ulberg LC. 1955. Synchronization of estrous cycles. Presented at Reprod. Fertil. Centen. Symp., Mich. State Univ., East Lansing 70. Hansel W, Malven PV. 1960. Estrous cycle regulation in beef cattle by orally active progestational agents. J. Anim. Sci. 19:1324



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71. Nellor JE, Ahrenhold JE, Nelson RH. 1960. Influence of oral administration of 6-methyl-17-acetoxyprogesterone on follicular growth and estrous behavior in beef heifers. J. Anim. Sci. 19:1331 72. Hansel W. 1966. Control of the ovarian cycle in cattle. In Reproduction in the Female Mammal: Proceedings of the Thirteenth Easter School in Agricultural Science, ed. GE Lamming, EC Amoroso, pp. 419–43. London: Butterworths 73. Robinson TJ. 1967. Control of the ovarian cycle in the sheep. In Reproduction in the Female Mammal: Proceedings of the Thirteenth Easter School in Agricultural Science, ed. GE Lamming, EC Amoroso, pp. 373–418. London: Butterworths 74. Hansel W, Schecter RE. 1972. Biotechnical procedures for control of the estrous cycles of domestic animals. Proc. VII Int. Congr. Anim. Reprod. Artif. Insemin., Munich, Germany, pp. 78–96 75. Hansel W, Beal WE. 1979. Ovulation control in cattle. In Beltsville Symposia in Agricultural Research Vol. 3: Animal Reproduction, ed. HW Hawk, pp. 91–109. Montclair, NJ: Allanheld, Osman 76. Smith RD, Pomerantz AJ, Beal WE, McCann JP, Pilbeam TE, Hansel W. 1984. Insemination of Holstein heifers at a preset time after estrous cycle synchronization using progesterone and prostaglandin. J. Anim. Sci. 58:792–800 77. Wilson DJ, Mallory DA, Busch DC, Leitman NR, Haden JK, et al. 2010. Comparison of short-term progestin-based protocols to synchronize estrus and ovulation in postpartum beef cows. J. Anim. Sci. 88:2045–54 78. Hansel W, Weir BJ. 1990. Genetic engineering of animals. J. Reprod. Fertil. S41:1–110 79. Thibodeaux JK, Del Vecchio RP, Hansel W. 1993. Role of platelet-derived growth factor in development of in vitro matured and in vitro fertilized bovine embryos. J. Reprod. Fertil. 98:61–66 80. Hansel W, Lim JM. 1998. In vitro fertilization and early embryo development. In Nutrition and Reproduction, ed. W Hansel, G Bray, DH Ryan, pp. 125–44. Baton Rouge: La. State Univ. Press 81. Lim JM, Hansel W. 1996. Roles of growth factors in the development of bovine embryos fertilized in vitro and cultured singly in a defined medium. Reprod. Fertil. Dev. 8:1199–205 82. Lim JM, Rocha A, Hansel W. 1996. A serum-free medium for use in a cumulus cell co-culture system for bovine embryos derived from in vitro maturation and in vitro fertilization. Theriogenology 45:1081–89 83. Lim JM, Hansel W. 1998. Improved development of in vitro–derived bovine embryos by use of a nitric oxide scavenger in a cumulus–granulosa cell coculture system. Mol. Reprod. Dev. 50:45–53 84. Lim JM, Mei Y, Chen B, Godke RA, Hansel W. 1999. Development of bovine IVF oocytes cultured in medium supplemented with a nitric oxide scavenger or inhibitor in a co-culture system. Theriogenology 51:941–49 85. Rocha A, Randel RD, Broussard JR, Lim JM, Blair RM, et al. 1998. High environmental temperature and humidity decrease oocyte quality in Bos taurus but not in Bos indicus cows. Theriogenology 49:657–65 86. Concannon PW, Hansel W, Visek WJ. 1975. The ovarian cycle of the bitch: plasma estrogen, LH and progesterone. Biol. Reprod. 13:112–21 87. Papke RL, Concannon PW, Travis HF, Hansel W. 1980. Control of luteal function and implantation in the mink by prolactin. J. Anim. Sci. 50:1102–7 88. Murphy BD, Concannon PW, Travis HF, Hansel W. 1981. Prolactin: the hypophyseal factor that terminates embryonic diapause in mink. Biol. Reprod. 25:487–91 89. Fernandez-Baca S, Hansel W, Novoa C. 1970. Corpus luteum function in the alpaca. Biol. Reprod. 3:252–61 90. Fernandez-Baca S, Hansel W, Saatman R, Sumar J, Novoa C. 1979. Differential luteolytic effects of right and left uterine horns in the alpaca. Biol. Reprod. 20:586–95 91. Rao CV. 1996. The beginning of a new era in reproductive biology and medicine: expression of low levels of functional luteinizing hormone/human chorionic gonadotropin receptors in nongonadal tissues. J. Physiol. Pharmacol. 47:41–53 92. Leuschner C, Hansel W. 2004. Membrane disrupting lytic peptides for cancer treatments. Curr. Pharm. Des. 10:2299–310 93. Hansel W, Leuschner C, Enright F. 2007. Destruction of breast cancers and their metastases by lytic peptide conjugates in vitro and in vivo. Mol. Cell. Endocrinol. 260–262:183–89

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94. Hansel W, Leuschner C, Gawronska B, Enright F. 2001. Targeted destruction of prostate cancer cells and xenografts by lytic peptide-bLH conjugates. Reprod. Biol. 1:20–32 95. Leuschner C, Enright FM, Melrose PA, Hansel W. 2001. Targeted destruction of androgen-sensitive and -insensitive prostate cancer cells and xenografts through luteinizing hormone receptors. Prostate 46:116– 25 96. Leuschner C, Enright FM, Gawronska-Kozak B, Hansel W. 2003. Human prostate cancer cells and xenografts are targeted and destroyed through luteinizing hormone releasing hormone receptors. Prostate 56:239–49 97. Gawronska B, Leuschner C, Enright FM, Hansel W. 2002. Effects of a lytic peptide conjugated to beta HCG on ovarian cancer: studies in vitro and in vivo. Gynecol. Oncol. 85:45–52 98. Leuschner C, Enright F, Gawronska B, Hansel W. 2003. Membrane disrupting lytic peptide conjugates destroy hormone dependent and independent breast cancer cells in vitro and in vivo. Breast Cancer Res. Treat. 78:17–27 99. Bodek G, Vierre S, Rivero-Muller A, Huhtaniemi I, Ziecik AJ, Rahman NA. 2005. A novel targeted therapy of Leydig and granulosa cell tumors through the luteinizing hormone receptor using a Hecatechorionic gonadotropin beta conjugate in transgenic mice. Neoplasia 7:497–508 100. Solipuram R, Aggarwal S, Leuschner C, Alila H, Hansel W. 2011. Pretreatment with FSH enhances the ability of EP-100 to target and destroy human pancreatic cells. Presented at Am. Assoc. Cancer Res. Symp., Orlando, FL 101. Hansel W, Enright F, Leuschner C. 2007. Conjugates of lytic peptides and LHRH or bCG target and cause necrosis of prostate cancers and metastases. Mol. Cell. Endocrinol. 269:26–33 102. Leuschner C, Hansel W. 2005. Targeting breast and prostate cancers through their hormone receptors. Biol. Reprod. 73:860–65 103. Aggarwal S, Ichikawa H, Takada Y, Sandur SK, Shishodia S, Aggarwal BB. 2006. Curcumin (diferuloylmethane) down-regulates expression of cell proliferation and antiapoptotic and metastatic gene products through suppression of IkBa kinase and Akt activation. Mol. Pharmacol. 69:195–206 104. Aggarwal S, Ndinguri MW, Solipuram R, Wakamatsu N, Hammer RP, et al. 2011. [DLys(6)]-luteinizing hormone releasing hormone-curcumin conjugate inhibits pancreatic cancer cell growth in vitro and in vivo. Int. J. Cancer 129:1611–23 105. Vuorenoja S, Rivero-Müller A, Ziecik AJ, Huhtaniemi I, Toppari J, Rahman NA. 2008. Targeted therapy for adrenocortical tumors in transgenic mice through their LH receptor by Hecate-human chorionic gonadotropin b conjugate. Endocr.-Relat. Cancer 15:635–48 106. Yates C, Sharp S, Jones J, Topps D, Coleman M, et al. 2011. LHRH-conjugated lytic peptides directly target prostate cancer cells. Biochem. Pharmacol. 81:104–10 107. Hansel W, McEntee K, Olafson P. 1951. The effects of two causative agents of experimental hyperkeratosis on vitamin A metabolism. Cornell Vet. 41:367–76 108. Hansel W, Olafson P, McEntee K. 1954. The isolation and identification of the causative agent of bovine hyperkeratosis (X-disease) from a processed wheat concentrate. Cornell Vet. 44:94–101 109. Barnhart KF, Christianson DR, Hanley PW, Driessen WH, Bernacky BJ, et al. 2011. A peptidomimetic targeting white fat causes weight loss and improved insulin resistance in obese monkeys. Sci. Transl. Med. 3:108ra12 110. Mullany LK, Richards JS. 2012. Minireview: animal models and mechanisms of ovarian cancer development. Endocrinology 153:1585–92 111. Trevino LS, Giles JR, Wang W, Urick ME, Johnson PA. 2010. Gene expression profiling reveals differentially expressed genes in ovarian cancer of the hen: Support for oviductal origin? Horm. Cancer 1:177–86 112. Hansel W. 1986. Animal agriculture for the year 2000 and beyond: William Henry Hatch Memorial Lecture. Presented for Natl. Assoc. State Univ. Land Grant Coll., U.S. Dept. Agric., Coop. State Res. Serv., Washington, DC 113. Hansel W, Godke RA. 1992. Future prospectives on animal biotechnology. Anim. Biotechnol. 3:111–37 114. Hansel W. 2003. The potential for improving the growth and development of cultured farm animal oocytes. Anim. Reprod. Sci. 79:191–201



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115. Paterson L, DeSousa P, Ritchie W, King T, Wilmut I. 2003. Application of reproductive biotechnology in animals: implications and potentials: applications of reproductive cloning. Anim. Reprod. Sci. 79:137– 43 116. Seidel GE Jr. 2003. Sexing mammalian sperm—intertwining of commerce, technology, and biology. Anim. Reprod. Sci. 79:145–56 117. Hasler JF. 2003. The current status and future of commercial embryo transfer in cattle. Anim. Reprod. Sci. 79:245–64 118. Albuz FK, Sasseville M, Lane M, Armstrong DT, Thompson JG, Gilchrist RB. 2010. Simulated physiological oocyte maturation (SPOM): a novel in vitro maturation system that substantially improves embryo yield and pregnancy outcomes. Hum. Reprod. 25:2999–3011 119. Wilmut I, Schnieke AE, McWhir J, Kind AJ, Campbell KH. 1997. Viable offspring derived from fetal and adult mammalian cells. Nature 385:810–13 120. Takahashi K, Yamanaka S. 2006. Induction of pluripotent stem cells from mouse embryonic and adult fibroblast cultures by defined factors. Cell 126:663–76 121. Takahashi K, Tanabe K, Ohnuki M. 2007. Induction of pluripotent stem cells from adult human fibroblasts by defined factors. Cell 131:861–72 122. Han X., Han J, Ding F, Cao S, Lim SS, et al. 2011. Generation of induced pluripotent stem cells from bovine embryonic fibroblast cells. Cell Res. 21:1509–12

20

Hansel

Annual Review of Animal Biosciences Volume 1, 2013

Contents


After 65 Years, Research Is Still Fun William Hansel . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1 Cross Talk Between Animal and Human Influenza Viruses Makoto Ozawa and Yoshihiro Kawaoka . . . . . . . . . . . . . . . . . . . . . . . . . 21 Porcine Circovirus Type 2 (PCV2): Pathogenesis and Interaction with the Immune System Xiang-Jin Meng . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 43 Evolution of B Cell Immunity David Parra, Fumio Takizawa, and J. Oriol Sunyer . . . . . . . . . . . . . . . . . . 65 Comparative Biology of gd T Cell Function in Humans, Mice, and Domestic Animals Jeff Holderness, Jodi F. Hedges, Andrew Ramstead, and Mark A. Jutila . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 99 Genetics of Pigmentation in Dogs and Cats Christopher B. Kaelin and Gregory S. Barsh . . . . . . . . . . . . . . . . . . . . . . 125 Cats: A Gold Mine for Ophthalmology Kristina Narfström, Koren Holland Deckman, and Marilyn Menotti-Raymond . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 157 Comparative Aspects of Mammary Gland Development and Homeostasis Anthony V. Capuco and Steven E. Ellis . . . . . . . . . . . . . . . . . . . . . . . . . . 179 Genetically Engineered Pig Models for Human Diseases Randall S. Prather, Monique Lorson, Jason W. Ross, Jeffrey J. Whyte, and Eric Walters . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 203 Accelerating Improvement of Livestock with Genomic Selection Theo Meuwissen, Ben Hayes, and Mike Goddard . . . . . . . . . . . . . . . . . . 221

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Integrated Genomic Approaches to Enhance Genetic Resistance in Chickens Hans H. Cheng, Pete Kaiser, and Susan J. Lamont . . . . . . . . . . . . . . . . . . 239 Conservation Genomics of Threatened Animal Species Cynthia C. Steiner, Andrea S. Putnam, Paquita E.A. Hoeck, and Oliver A. Ryder . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 261


Phytase, A New Life for an “Old” Enzyme Xin Gen Lei, Jeremy D. Weaver, Edward Mullaney, Abul H. Ullah, and Michael J. Azain . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 283 Effects of Heat Stress on Post-Absorptive Metabolism and Energetics Lance H. Baumgard and Robert P. Rhoads Jr. . . . . . . . . . . . . . . . . . . . . 311 Epigenetics: Setting Up Lifetime Production of Cows by Managing Nutrition R.N. Funston and A.F. Summers . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 339 Systems Physiology in Dairy Cattle: Nutritional Genomics and Beyond Juan J. Loor, Massimo Bionaz, and James K. Drackley . . . . . . . . . . . . . . 365 In Vivo and In Vitro Environmental Effects on Mammalian Oocyte Quality Rebecca L. Krisher . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 393 The Equine Endometrial Cup Reaction: A Fetomaternal Signal of Significance D.F. Antczak, Amanda M. de Mestre, Sandra Wilsher, and W.R. Allen . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 419 The Evolution of Epitheliochorial Placentation Anthony M. Carter and Allen C. Enders . . . . . . . . . . . . . . . . . . . . . . . . . 443 The Role of Productivity in Improving the Environmental Sustainability of Ruminant Production Systems Judith L. Capper and Dale E. Bauman . . . . . . . . . . . . . . . . . . . . . . . . . . 469 Making Slaughterhouses More Humane for Cattle, Pigs, and Sheep Temple Grandin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 491

Contents

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25 Years of FUN!

Still delirious after all these years.

Still Enthusiastic After All These Years.

Thirty Years of Magnetic Stimulation: Is it Still Only for the Purpose of Research?

Misdiagnosis in JME: Still a problem after 17 years?

After 10 years of AIDS, safe-sex message still controversial.

Isolated coronary artery bypass grafting in extracorporeal circulation in patients over 65 years old - does age still matter?

Fun and games in Berkeley: the early years (1956-2013).

My 65 years in protein chemistry.

Physicians after 65.

Data-driven, evidenced-based, computational modeling research is still needed in trauma research.

RNA World research-still evolving.

1958-2014: after 56 years of research, cytochrome p450 reactivity is finally explained.

Totally implantable vascular access devices 30 years after the first procedure. What has changed and what is still unsolved?

Comparison of outcomes after heart replacement therapy in patients over 65 years old.

Increased mortality after lower extremity fractures in patients <65 years of age.

Histopathological evaluation is still useful after bladder neck resection.

19 years outcome after cementless total hip arthroplasty with spongy metal structured implants in patients younger than 65 years.

Higgs inflation is still alive after the results from BICEP2.

Elective operation after acute complicated diverticulitis: is it still mandatory?

Long-term survival after surgical aortic valve replacement among patients over 65 years of age.

[Results after internal fixation of humerus distal fractures in patients over than 65 years old].

Visceral obesity is not an independent risk factor of mortality in subjects over 65 years.

5000 years old and still going strong.