Experimental Parasitology 143 (2014) 48–54

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Insights into regulatory molecules of intestinal epithelial cell turnover during experimental infection by Heterophyes heterophyes Dalia S. Ashour a, Ahmad A. Othman a,⇑, Dina A. Radi b a b

Department of Medical Parasitology, Faculty of Medicine, Tanta University, Tanta, Egypt Department of Pathology, Faculty of Medicine, Tanta University, Tanta, Egypt

h i g h l i g h t s

g r a p h i c a l a b s t r a c t

 Heterophyes induces structural and

Heterophyes heterophyes adult embedded in between the villi with inflammation and apoptosis of cells.

inflammatory alterations in the small intestinal mucosa.  Caspase-3 increases in intestinal cells of infected animals during early heterophysiasis.  NF-jB increases over time in infected animals, denoting a possible role in intestinal cell survival.

a r t i c l e

i n f o

Article history: Received 2 December 2013 Received in revised form 27 April 2014 Accepted 11 May 2014 Available online 19 May 2014 Keywords: Heterophyes heterophyes Caspase-3 Nuclear factor-jB Intestinal cell Apoptosis

a b s t r a c t Heterophyiasis is an intestinal disease that remains endemic in many parts of the world, particularly the Nile Delta of Egypt and Southeast Asia, yet the populations at risk of infection expand throughout the world. The main histopathological feature of infection is villous atrophy, but the underlying factors are not well understood. Apoptosis of the villous epithelial cells was previously reported to be enhanced during intestinal parasitic infections; however, the role of Heterophyes heterophyes on enterocyte apoptosis was to be explored. Therefore, intestinal sections from mice experimentally infected with H. heterophyes were studied histopathologically and immunohistochemically for caspase-3 and NF-jB and compared to non-infected control mice. Atrophic villi covered by poorly differentiated epithelial cells were observed in the 2nd week post-infection. Also, we noted marked hyperplasia of the intestinal crypts with abundant inflammatory cellular infiltrate in the lamina propria, as well as apoptosis of cells lining the intestinal villi. Both caspase-3 and NF-jB showed positive staining in the intestinal epithelial cells with varying grades of intensity over the length of infection. Caspase-3 expression rose at the 2nd week p.i. then decreased over time, whereas NF-jB expression showed progressive increase throughout the weeks of infection. In conclusion, caspase-3 activation may be an important factor in the apoptotic pathway in early heterophyiasis, and, on the other hand, NF-jB seems to play a role in protecting the intestinal cells from excessive apoptosis. These observations may help open new avenues for tissue protective therapies that avoid or control the deleterious processes of apoptosis in various inflammatory conditions. Ó 2014 Elsevier Inc. All rights reserved.

1. Introduction ⇑ Corresponding author. Address: Parasitology Department, Tanta Faculty of Medicine, Tanta, Gharbiya, Egypt. E-mail address: [email protected] (A.A. Othman). http://dx.doi.org/10.1016/j.exppara.2014.05.003 0014-4894/Ó 2014 Elsevier Inc. All rights reserved.

In global terms, intestinal trematodiases are the commonest parasitic infections in humans and animals. Although less associated

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with mortality than other groups of parasites, they are responsible for significant morbidity (Toledo et al., 2006). Heterophyiasis is an intestinal illness caused by infection with the heterophyid digenetic flukes. The disease is still endemic in many parts of the world, particularly the Nile Delta of Egypt and Southeast Asia, where favorable conditions exist for parasite propagation including the availability and abundance of the intermediate hosts, increasing production of fish in unhygienic conditions due to frequent disposal of human wastes directly into rivers and lakes, and the habit of consuming raw or inadequately processed fish (Abou-Basha et al., 2000; Graczyk and Fried, 2007). Indeed, the populations at risk and the geographical range of the infection expand because of the rapid growth of fish industry, improved transportation systems, and demographic changes such as population movements (Chai et al., 2005). Humans become infected while eating fish harboring viable metacercariae that mature into adults within 5–10 days. Most infections of heterophyids are asymptomatic or accompanied by mild intestinal discomfort and mucous diarrhea (Toledo et al., 2006; Elsheikha, 2007). However, the eggs (and sometimes adult worms) may then enter nearby lymphatics or blood vessels and be transported to other organs, such as the heart, spinal cord or brain, lungs, liver and spleen (Chai and Lee, 2002). The main histopathological features in the early phase of infection with Heterophyidae include villous atrophy, characterized by fusion, thickening, and shortening of the villi; hypertrophy of the crypts of Lieberkühn, resulting in a decrease of the villus/crypt ratio; enlargement of mesenteric lymph nodes; and inflammatory responses with cell infiltration (Hong et al., 1997; Yu et al., 1997). In this context, the factors underlying villous atrophy in the related species Metagonimus yokogawai are not well understood. One mechanism may be mechanical damage to the epithelial cells by the attached worms. However, other factors appear to be involved (Toledo et al., 2006). Yu et al. (1997) suggested that M. yokogawai induces villous atrophy via inhibition of cell proliferation in the intestinal crypts. Apoptosis of the villous epithelial cells was previously reported to be enhanced during intestinal parasitic infections which may be relevant to rapid removal of damaged cells (Kuroda et al., 2002). Caspase-3 is the core member of caspase family. It is a well-characterized apoptotic effector. When activated, it can cleave the vast majority of polypeptides that undergo proteolysis in apoptotic cells (Park et al., 2001; Ben-Yehudah et al., 2003). On the other hand, nuclear factor kappa B (NF-jB) is a key regulator of inflammation and innate immunity (Ghosh and Karin, 2002). It maintains tissue integrity and might play an anti-inflammatory role. Moreover, NF-jB protects cells from death by inducing expression of antiapoptotic proteins (Karin and Lin, 2002; Pasparakis, 2009). The role of Heterophyes heterophyes (H. heterophyes) upon induction of enterocyte apoptosis still remains to be elucidated. Therefore, based on the transient and self limiting manifestations of the infection, we hypothesize that Heterophyidae might have evolved means to control host cell apoptosis in order to facilitate the establishment of infection in the newly exposed host. 2. Materials and methods

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2.2. Animals and experimental design The current study included 60 male Swiss albino mice, four to six weeks old and weighing 18–20 g. Mice were maintained and infected in accordance with the institutional and national guidelines. Mice were divided into two groups: group (I), 15 noninfected mice (control group), and group (II), 45 mice that were infected with encysted metacercariae of H. heterophyes. Mice were infected by receiving 300 encysted metacercariae each orally after overnight fasting. The metacercariae were introduced into their stomach by using one milliliter insulin syringe fitted with 18 gauge blunt ended needle. Thereafter, groups of 15 infected mice were sacrificed at the end of the 2nd, 3rd, and 4th weeks post-infection (p.i.). Control mice were sacrificed at the second week of the experiment. The middle thirds of the small intestines from all groups were taken for histopathological and immunohistochemical studies.

2.3. Histopathological and immunohistochemical studies Histopathological examination was performed on tissue specimens fixed in 10% formalin, washed, dehydrated in ascending grades of alcohol, cleared in xylene, embedded in paraffin wax, sectioned and stained with hematoxylin and eosin (H&E). For assessment of counts of various inflammatory cells, goblet cells, and apoptotic cells per high power fields (HPF) of the examined intestinal sections, a semiquantitative score was used as follows: 0: Nil, 1: Few (up to 10 cells/HPF), 2: Moderate number (11–40 cells/ HPF), and 3: Numerous (more than 40 cells/HPF). For immunohistochemical staining for caspase-3 and NF-jB, sections were mounted on poly-L-lysine-coated slides. The avidin–biotin–peroxidase method was performed using primary monoclonal antibody against caspase-3 (‘‘Clone 3CSP01’’, Readyto-use, Kit No. RB-1197-B, Lab Vision, CA), or primary monoclonal antibody against NF-jB (IgG isotype, molecular weight 65 kDa, diluted at 1:1000, Kit No. MS-1126-P0, Lab Vision, CA), respectively. Briefly, the sections were deparafinized and endogenous peroxidase activity was blocked using a 0.3% solution of hydrogen peroxidase in phosphate buffered saline (PBS) at room temperature for 10 min. After microwave treatment, primary antibody was applied for 30 min at room temperature and washed in PBS. Linking antibody and streptavidin–peroxidase complex (Lab Vision, CA) were added consecutively for 10 min at room temperature and washed in PBS. The peroxidase activity was visualized with diaminobenzidine (Lab Vision, CA) applied for 5 min. Appropriate positive and negative controls were also labeled with the primary antibody. Caspase-3 and NF-jB immune stains were assessed microscopically in HPF under 400 magnification by examining 20 HPF areas of small intestinal sections of each examined animal. Positive cells for caspase-3 immune stain show brownish cytoplasmic and some nuclear staining with variable intensities graded as +1 (weak), +2 (moderate) and +3 (strong). Positive cells for NF-jB immune stain show brownish cytoplasmic staining with variable intensities graded as +1 (weak), +2 (moderate) and +3 (strong).

2.1. Collection of H. heterophyes metacercariae 2.4. Statistical analysis The encysted metacercariae of H. heterophyes were collected from Tilapia species of fish, from Dakahlia Governorate brackish water lakes (Manzalla), Egypt, by pepsin digestion method as previously described by Yokogawa and Sano (1968). The final volume of metacercarial suspension was adjusted by saline to contain about 300 encysted metacercariae per 0.5 ml. (the infective dose).

Data were presented as mean ± standard deviation. The probability of significant differences among means of groups was determined by Chi-square test using Statistical Package of Social Sciences (SPSS) software for windows, version 16. P < 0.05 was considered statistically significant.

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3. Results 3.1. Histopathological changes in the small intestine Histopathological examination of the small intestinal sections of control mice revealed normal intestinal tissue with a negligible number of neutrophils and lympocytes, absent eosinophils, and normal goblet cells. In contrast, the sections of the infected group showed inflammatory reaction especially near the areas of attachment of the parasite with notable dynamic changes over the weeks of infection. At 2 weeks p.i., the villi were shorter and blunter than normal; the atrophic villi were covered by poorly differentiated lower columnar epithelial cells, and there was fusion of the lateral surfaces and tips of villi in some areas, with mild erosion of the surface of villi (Fig. 1A). Also, the pathological alterations included marked and widespread hyperplasia of the intestinal crypts (Fig. 1B); goblet cell hyperplasia; massive inflammatory cellular infiltrate in the lamina propria composed of eosinophils, neutrophils, lymphocytes, and some histiocytes (Fig. 1C); apoptosis of cells lining the intestinal villi manifested by dark stained nuclei due to clumped chromatin (Fig. 1D); and adult Heterophyes worms in between some intestinal villi (Fig. 1E). Later at 3 weeks p.i., the examination of intestinal sections showed that the number of acute inflammatory cells (neutophils and eosinophils) started to decline, while chronic inflammatory

cells (lymphocytes and histiocytes) were relatively increased in number. Also lesser degree of goblet cell hyperplasia was found as the infection subsided gradually. Apoptotic cells were also less abundant (Fig. 1F). At 4 weeks p.i., the examined sections revealed almost normal histopathological findings with few mononuclear inflammatory cells. The overall dynamic changes of the inflammatory cellular infiltrate, goblet cells, and apoptotic cells over the weeks of infection were depicted in (Table 1). 3.2. Immunohistochemical study On examination of the caspase-3 immunostained sections, none of control cases (0%) were positive for caspase-3 expression. Among the infected group 2 weeks p.i., all cases (100%) were positive for caspase-3 expression with variable intensity, where 6 out of 15 cases showed moderate caspase-3 expression (+2), and the remaining 9 cases showed strong caspase-3 expression (+3) (Fig. 2A and B). At 3 weeks p.i., caspase-3 expression was noticed in 12 out of 15 cases (80%) including 3 cases with strong caspase-3 expression (+3), 6 cases with moderate caspase-3 expression (+2) (Fig. 2C) and 3 cases with weak caspase-3 expression (+1), while the remaining 3 cases (20%) were negative for caspase-3 expression. On the other hand, the infected mice 4 weeks p.i. showed caspase-3 expression in 9 out of 15 cases (60%) including 3 cases showing moderate expression (+2), and 6 cases with

Fig. 1. Histopathological changes in the small intestine of mice infected with H. heterophyes at 2 weeks (A–E) and 3 weeks (F) p.i.: (A) there are villous atrophy, fusion of adjacent villi, and inflammatory cell infiltration, as well as vascular dilatation, congestion and edema in the villous interstitium and lamina propria (100), (B) the intestinal crypts show marked proliferation (200), (C) marked local inflammatory cellular infiltrate with lymphocytes (white arrows) and eosinophils (black arrows) is noted (200), (D) dark-stained nuclei of many intestinal cells (arrows) denoting nuclear pyknosis are observed (200), (E) adult H. heterophyes (arrow) is embedded in between the villi (200), and (F) moderate inflammatory cellular infiltrate in the lamina propria is seen (200).

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D.S. Ashour et al. / Experimental Parasitology 143 (2014) 48–54 Table 1 Semiquantitative score of cells of inflammation, goblet cells, and apoptotic cells.

Control 2 weeks p.i. 3 weeks p.i. 4 weeks p.i.

Neutrophils

Eosinophils

Lymphocytes

Histiocytes

Goblet cells

Apoptotic cells

1 3 1 0

0 3 2 1

1 2 3 1

0 2 3 1

1 3 2 1

1 3 2 1

N.B. 0: nil, 1: few (up to 10 cells/HPF), 2: moderate number (11–40 cells/HPF), and 3: numerous (more than 40 cells/HPF).

Fig. 2. Immunostaining for caspase-3 in the epithelial cells lining intestinal villi of mice infected with H. heterophyes: (A) at 2 weeks p.i., the epithelial cells show moderate cytoplasmic expression (400), (B) other cases, at 2 weeks p.i., show a strong expression (400), (C) at 3 weeks p.i., the cells show moderate cytoplasmic immune staining (400), and (D) at 4 weeks p.i., weak cytoplasmic immune staining is seen (400).

weak expression (+1) (Fig. 2D), while the remaining 6 cases (40%) were completely negative for caspase-3 expression. Although caspase-3 expression was decreasing over time, significant correlations were found between the infected group at different time points (2, 3, and 4 weeks p.i.) as compared to the control group (P < 0.01, 0.02, and 0.03, respectively) (Table 2). Examination of the sections immunostained for NF-jB revealed that 3 out of 15 cases (20%) of control mice showed weak expression (+1) in the cytoplasm of epithelial cells lining the intestinal villi, while the remaining 12 cases (80%) were completely negative for NF-jB expression. Infected mice at 2 weeks p.i. showed positive cytoplasmic expression for NF-jB in all cases (100%) with variable intensities: 6 cases showed moderate expression (+2) and 9 cases showed weak cytoplasmic expression for NF-jB (Fig. 3A). All cases (100%) of infected mice 3 weeks p.i. were positive for NF-jB

expression, including 6 cases showing moderate cytoplasmic expression (+2) (Fig. 3B), 6 cases with strong cytoplasmic expression (+3), and 3 cases showing weak cytoplasmic expression for NF-jB. Regarding infected mice 4 weeks p.i., all cases were also positive for NF-jB cytoplasmic expression where 9 cases showed strong cytoplasmic expression (+3) (Fig. 3C), and 6 cases showed moderate expression for NF-jB (+2). Expression of NF-jB in the infected group at all time points of infection (2, 3, and 4 weeks) was noted to be significantly increasing over the weeks post-infection (P < 0.04, 0.03, and 0.02, respectively) (Table 2) as compared to the control group. As regards the trend of expression of caspase-3 and NF-jB among all the studied groups, it was found that caspase-3 expression surged early in the infection then declined with time, whereas NF-jB expression was increasing gradually over the successive weeks of infection (Fig. 4).

Table 2 Score (mean ± SD) of caspase-3 and NF-jB immunoreactivity.

Caspase-3 NF-jB *

Control

2 weeks p.i.

3 weeks p.i.

4 weeks p.i.

0 0.2 ± 0.447

2.6 ± 0.548* 1.4 ± 0.548*

1.6 ± 1.140* 2.2 ± 0.837*

0.8 ± 0.837* 2.6 ± 0.547*

Statistically significant difference (P < 0.05) as compared to control group.

4. Discussion Apoptosis is a form of programmed cell death involved in a wide range of adaptive processes, from embryogenesis to stress injury responses. Detrimental effects caused by apoptosis can be

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Fig. 3. Immunostaining for NF-jB in the epithelial cells lining intestinal villi of mice infected with H. heterophyes showing (A) weak cytoplasmic immune staining at 2 weeks p.i. (200), (B) moderate cytoplasmic expression at 3 weeks p.i. (400), and (C) strong nuclear and cytoplasmic immune staining at 4 weeks p.i. (200).

Score of immunostain

3 2.5 2 1.5

Caspase 3 NF Kappa B

1 0.5 0

C

2w

3w

4w

Groups Fig. 4. Caspase-3 and NF-jB expression in different durations of infection.

triggered by parasitic infections, depending upon the specific host– parasite situation. During their evolution, parasites have developed mechanisms to induce or avoid host cell apoptosis in order to be able to survive and complete their life cycle (Lawen, 2003). Certain inflammatory disorders of the small intestine including parasitic infections are associated with enhanced apoptosis in the intestinal epithelial cells (IEC) with crypt hyperplasia; the latter reflecting enhanced replacement of damaged IEC (Hyoh et al., 1999). Histopathological and immunohistochemical studies (caspase-3 and NF-jB) were performed in the current study to explain the pathological changes in intestinal cells during experimental infection by H. heterophyes. Our results demonstrated that H. heterophyes induced apoptosis in small intestinal epithelial cells early in the infection and that the apoptotic activity declined over the next weeks p.i. as evidenced by the histopathological findings and the inverse relationship of caspase-3 and NF-jB expression in the intestinal tissues. The current study of experimental H. heterophyes infection reports histopathological changes in the intestine that include

massive hyperplasia of the intestinal crypts with inflammatory cellular infiltrate in the lamina propria, and apoptosis of cells lining the intestinal villi. Similar findings were described in Heterophyidae. Wattanagoon and Bunnag (2013) indicated that the flukes attach to the mucosa of the small intestine and may cause mild inflammation with cellular infiltration, shallow ulcers, local necrosis and hemorrhage. The underlying submucosa is slightly thickened by edema and little cellular infiltration. In Haplorchis taichui infection, the histopathological findings include mucosal ulceration; mucosal and submucosal hemorrhages; atrophy, fusion and shortening of the villi; chronic inflammation; and fibrosis of the submucosa with eosinophil and lymphocyte infiltration (Sukontason et al., 2005). Apoptotic cells are normally observed at the tips of villi as well as in crypts (Hall et al., 1994). In certain pathological inflammatory conditions such as coeliac disease and nematode infections, numbers of apoptotic nuclei were reported to be increased in villus epithelial cells, indicating that apoptosis has important roles not only in physiological replacement of villous epithelial cells but also in pathological conditions (Moss et al., 1996; Hyoh et al., 1999). In the current study, the caspase-3 stained sections showed positive reaction in cellular lining of infected intestinal villi in 100% of cases 2 weeks p.i., in 80% of cases 3 weeks p.i., and in 60% of cases 4 weeks p.i., with variable degrees of staining intensity. Activation of caspase-3 in the infected cells indicates that apoptosis was in action in these cells. Apoptosis is associated with activation of effector molecules, including the key protease caspase-3 (van der Woude et al., 2003; Pan et al., 2003). Caspases are a group of enzymes that are present in cells as inactive zymogens that are activated by proteolytic cleavage upon cellular stimulation (Li and Yuan, 2008). Oxidative stress could be responsible for apoptosis by activating caspase and triggering the apoptotic cascade (Yaguchi et al., 2010). Caspase-3 activates the DNA-degrading endonuclease CAD (caspase-activated DNase) responsible for DNA degradation leading to eventual cell death (Duprez et al., 2009).

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Apoptosis leads to the packaging of cell contents into apoptotic bodies that can be phagocytosed by other cells without induction of a strong inflammatory response (Weinrauch and Zychlinsky, 1999; Heussler et al., 2001). Apoptosis of infected host cells is widely recognized as an immediate defense response to limit the spread of certain infections (Heussler et al., 2001; Green, 2005). Therefore, the initiation of apoptosis in parasite-infected host cells could be beneficial for the host, both limiting the number of adherent parasites and overwhelming the inflammatory responses (Oberhuber et al., 1997). This could explain the increase of caspase-3 expression and apoptosis early in H. heterophyes infection. In addition, apoptosis ensures the integrity of the epithelial barrier and prevents unwarranted activation of the inflammatory responses (Liu et al., 2008). It was found that Nippostrongylus brasiliensis induced significant increases in caspase-3 activity in IEC which may reflect accelerated apoptosis in IEC after infection (Hyoh et al., 2002). Similar changes were observed with Cryptosporidium parvum (Chen et al., 1999) and Trichinella spiralis (Karman´ska et al., 2000). However, excess apoptosis of enterocytes may contribute to the pathogenesis by accelerating the normal turnover of the intestinal epithelium and disrupting epithelial barrier function as in giardiasis. This, in turn, may cause electrolyte and nutrient malabsorption (Panaro et al., 2007). That is why many intracellular pathogens target caspase signaling as a means to impede apoptosis of the infected host cell. Apparently, inhibition of caspases likely contributes to host cell survival. Therefore, unsurprisingly, C. parvum inhibits apoptosis of infected human small intestinal epithelial cells (Liu et al., 2008). NF-jB is a key regulator of inflammation and innate immunity (Ghosh and Karin, 2002). It is sequestered in the cytoplasm of resting cells by the IjB family of proteins. Following exposure of cells to infectious or inflammatory stimuli, IjB is phosphorylated and degraded, allowing NF-jB to translocate to the nucleus and influence the transcription of a wide range of immune response genes (Ghosh, 1999; Karin and Ben-Neriah, 2000). NF-jB transcription factors activate expression of genes encoding cytokines, chemokines, and adhesion molecules (Lawrence, 2009; Pasparakis, 2009). Through such genes, NF-jB plays a key role in coordination of inflammation and immune responses. NF-jB could also maintain tissue integrity and protect cells from death by inducing expression of anti-apoptotic proteins (Karin and Lin, 2002; Pasparakis, 2009). Moreover, NF-jB is critical in regulating inflammation in the intestine. Activation of NF-jB and expression of inflammatory cytokines has been reported in human and murine models of inflammatory bowel disease (Neurath et al., 1998). Cellular protection by NF-jB is in part mediated by its direct effects on epithelial cell survival (Greten et al., 2004). Research suggests that NF-jB protects against cell damage in a paracrine manner by up-regulating the expression of chemokines that recruit myeloid and T cells (Eckmann et al., 2008). However, NF-jBdependent cell survival and chemokine expression is dominant in epithelial cells under acute injury conditions, whereas proinflammatory functions of NF-jB are decisive in chronic situations of immune dysregulation (Eckmann et al., 2008). In the present study, sections stained with NF-jB showed positive reaction in 100% of all cases of the infected groups with increasing intensity of staining over time. Similarly, it has been shown that NF-jB activation is associated with C. parvum infection of epithelial cells and that inhibiting NF-jB activation promotes apoptosis (Chen et al., 2001). Caspases can act as inhibitors of NF-jB activity. Several proteins throughout the NF-jB signaling cascades are cleaved by caspases in response to inducers of programmed cell death, thus leading to impaired NF-jB activation (Frelin et al., 2008; Tang et al., 2001). In addition, Ravi et al. (1998) found that CD95, which is known as Fas, possesses an apoptotic effect, and can stimulate

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NF-jB degradation by caspases. Additionally, when an antibody against CD95 was used, caspases were inhibited and the inducibility of NF-jB was restored. This study suggests that H. heterophyes inhibits apoptosis after a short period of caspase-3 activation. The same trend was observed in leishmaniasis. Apoptosis is inhibited in the presence of viable Leishmania infantum promastigotes in culture medium via a mechanism involving inhibition of caspase-3 activation (Lisi et al., 2005). In conclusion, this study has provided a preliminary description of the changes of enterocyte’s apoptotic pathways during experimental infection by H. heterophyes. It seems that caspase-3 activation is an important factor in apoptosis in early heterophyiasis. Furthermore, there is a linkage between NF-jB signaling process and the apoptotic changes in the infected intestinal cells, supporting the role of NF-jB in protecting the intestinal cells from excessive apoptosis. The balance between caspase-3 and NF-jB seems to determine the fate of IEC. Whether these specific changes were triggered or favored by parasite factors awaits further research. Finally, these findings provide some lines of evidence that the apoptotic process could be reversed by anti-apoptotic agents leading to the opportunity to rescue cells. These observations may open new avenues for tissue protective therapies that avoid or control various deleterious processes of apoptosis. References Abou-Basha, L.M., Abdel-Fattah, M., Orecchia, P., Dicave, D., Zaki, A., 2000. Epidemological study of heterophyiasis among humans in an area of Egypt. East. Mediterr. Health J. 6, 932–938. Ben-Yehudah, A., Aqeilan, R., Robashkevich, D., Lorberboum-Galski, H., 2003. Using apoptosis for targeted cancer therapy by a new gonadotropin releasing hormone-DNA fragmentation factor 40 chimeric protein. Clin. Cancer Res. 9, 1179–1190. Chai, J.Y., Lee, S.H., 2002. Food-borne intestinal trematode infections in the Republic of Korea. Parasitol. Int. 51, 129–154. Chai, J.Y., Murrell, K.D., Lymbery, A.J., 2005. Fish borne parasitic zoonoses: status and issues. Int. J. Parasitol. 35, 1233–1254. Chen, X.M., Gores, G.J., Paya, C.V., La Russo, N.F., 1999. Cryptosporidium parvum induces apoptosis in biliary epithelia by a Fas/Fas ligand-dependent mechanism. Am. J. Physiol. 277, 599–608. Chen, X.M., Levine, S.A., Splinter, P.L., Tietz, P.S., Ganong, A.L., Jobin, C., Gores, G.J., Paya, C.V., La Russo, N.F., 2001. Cryptosporidium parvum activates nuclear factor kappa B in biliary epithelia preventing epithelial cell apoptosis. Gastroenterology 120, 1774–1783. Duprez, L., Wirawan, E., Vanden Berghe, T., Vandenabeele, P., 2009. Major cell death pathways at a glance. Microbes Infect. 11, 1050–1062. Eckmann, L., Nebelsiek, T., Fingerle, A.A., Dann, S.M., Mages, J., Lang, R., Robine, S., Kagnoff, M.F., Schmid, R.M., Karin, M., Arkan, M.C., Greten, F.R., 2008. Opposing functions of IKK beta during acute and chronic intestinal inflammation. Proc. Natl. Acad. Sci. U.S.A. 105 (39), 15058–15063. Elsheikha, H.M., 2007. Heterophyosis: risk of ectopic infection. Vet. Parasitol. 147 (3–4), 341–342. Frelin, C., Imbert, V., Bottero, V., Gonthier, N., Samraj, A.K., Schulze-Osthoff, K., Auberger, P., Courtois, G., Peyron, J.F., 2008. Inhibition of the NF-kappa B survival pathway via caspase-dependent cleavage of the IKK complex scaffold protein and NF-kappa B essential modulator NEMO. Cell Death Differ. 15, 152–160. Ghosh, S., 1999. Regulation of inducible gene expression by the transcription factor NF-jB. Immunol. Res. 19, 183–189. Ghosh, S., Karin, M., 2002. Missing pieces in the NF-kappa B puzzle. Cell 109 (Suppl.), S81–S96. Graczyk, T.K., Fried, B., 2007. Human waterborne trematode and protozoan infections. Adv. Parasitol. 64, 111–160. Green, D.R., 2005. Apoptotic pathways: ten minutes to dead. Cell 121, 671–674. Greten, F.R., Eckmann, L., Greten, T.F., Park, J.M., Li, Z.W., Egan, L.J., Kagnoff, M.F., Karin, M., 2004. IKK beta links inflammation and tumorigenesis in a mouse model of colitis-associated cancer. Cell 118, 285–296. Hall, P.A., Coates, P.J., Ansari, B., et al., 1994. Regulation of cell number in the mammalian gastrointestinal tract: the importance of apoptosis. J. Cell Sci. 107, 3569–3577. Heussler, V.T., Kuenzi, P., Rottenberg, S., 2001. Inhibition of apoptosis by intracellular protozoan parasites. Int. J. Parasitol. 31, 1166–1176. Hong, S.J., Han, J.H., Park, C.K., Kang, S.Y., 1997. Intestinal pathologic findings at early stage infection by Centrocestus armatus in albino rats. Korean J. Parasitol. 35, 135–138. Hyoh, Y., Ishizaka, S., Horii, T., Fujiwara, A., Tegoshi, T., Yamada, M., Arizono, N., 2002. Activation of caspases in intestinal villus epithelial cells of normal and nematode infected rats. Gut 50, 71–77.

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Hyoh, Y., Nishida, M., Tegoshi, T., et al., 1999. Enhancement of apoptosis with loss of cellular adherence in the villus epithelium of the small intestine after infection with the nematode Nippostrongylus brasiliensis in rats. Parasitology 119, 199–207. Karin, M., Ben-Neriah, Y., 2000. Phosphorylation meets ubiquitination: the control of NF-jB activity. Annu. Rev. Immunol. 18, 621–623. Karin, M., Lin, A., 2002. NF-kappa B at the crossroads of life and death. Nat. Immunol. 3, 221–227. Karman´ska, K., Houszka, M., Piekarska, J., 2000. The phenomenon of apoptosis in the course of experimental trichinellosis in mice. Wiad. Parazytolog. 46, 111–115. Kuroda, A., Uchikawa, R., Matsuda, S., Yamada, M., Tegoshi, T., Arizono, N., 2002. Upregulation of Fas (CD95) and induction of apoptosis in intestinal epithelial cells by nematode-derived molecules. Infect. Immun. 70 (8), 4002–4008. Lawen, A., 2003. Apoptosis: an introduction. BioEssays 25, 888–897. Lawrence, T., 2009. The nuclear factor NF-kappa B pathway in inflammation. Cold Spring Harb. Perspect. Biol. 1 (6), a001651. Li, J., Yuan, J., 2008. Caspases in apoptosis and beyond. Oncogene 27, 6194–6206. Lisi, S., Sisto, M., Acquafredda, A., Spinelli, R., Schiavone, M., Mitolo, V., Brandonisio, O., Panaro, M., 2005. Infection with Leishmania infantum inhibits actinomycin Dinduced apoptosis of human monocytic cell line U-937. J. Eukaryot. Microbiol. 52, 211–217. Liu, J., Enomoto, S., Lancto, C.A., Abrahamsen, M.S., Rutherford, M.S., 2008. Inhibition of apoptosis in Cryptosporidium parvum-infected intestinal epithelial cells is dependent on survivin. Infect. Immun. 76 (8), 3784–3792. Moss, S.F., Attia, L., Scholes, J.V., et al., 1996. Increased small intestinal apoptosis in coeliac disease. Gut 39, 811–817. Neurath, M.F., Fuss, I., Schurmann, G., Pettersson, S., Arnold, K., Muller-Lobeck, H., Strober, W., Herfarth, C., Buschenfelde, K.H., 1998. Cytokine gene transcription by NF-jB family members in patients with inflammatory bowel disease. Ann. N.Y. Acad. Sci. 859, 149–159. Oberhuber, G., Kastner, N., Stolte, M., 1997. Giardiasis: a histologic analysis of 567 cases. Scand. J. Gastroenterol. 32, 48–51. Pan, W., Ishii, H.H., Ebihara, Y., Grobe, G.C., 2003. Prognostic use of growth characteristics of early gastric cancer and expression patterns of apoptotic, cell proliferation, and cell adhesion proteins. J. Surg. Oncol. 82, 104–110. Panaro, M.A., Cianciulli, A., Mitolo, V., Mitolo, C.I., Acquafredda, A., Brandonisio, O., Cavallo, P., 2007. Caspase-dependent apoptosis of the HCT-8 epithelial cell line

induced by the parasite Giardia intestinalis. FEMS Immunol. Med. Microbiol. 51 (2), 302–309. Park, I.C., Park, M.J., Rhee, C.H., et al., 2001. Protein kinase C activation by PMA rapidly induces apoptosis through caspase-3/CPP32 and serine protease (s) in a gastric cancer cell line. Int. J. Oncol. 18, 1077–1083. Pasparakis, M., 2009. Regulation of tissue homeostasis by NF-kappa B signalling: implications for inflammatory diseases. Nat. Rev. Immunol. 9, 778–788. Ravi, R., Bedi, A., Fuchs, E.J., Bedi, A., 1998. CD95 (Fas)-induced caspase-mediated proteolysis of NF-jB. Cancer Res. 58, 882–886. Sukontason, K., Unpunyo, P., Sukontason, K.L., Piangjai, S., 2005. Evidence of Haplorchis taichui infection as pathogenic parasite: three case reports. Scand. J. Infect. Dis. 37, 388–390. Tang, G., Yang, J., Minemoto, Y., Lin, A., 2001. Blocking caspase-3-mediated proteolysis of IKK beta suppresses TNF-alpha-induced apoptosis. Mol. Cell 8, 1005–1016. Toledo, R., Esteban, J.G., Fried, B., 2006. Immunology and pathology of intestinal trematodes in their definitive hosts. Adv. Parasitol. 63, 285–365. van der Woude, C.J., Kleibeuker, J.H., Tiebosch, A.T., Homan, M., Beuving, A., Jansen, P.L., et al., 2003. Diffuse and intestinal type gastric carcinomas differ in their expression of apoptosis related proteins. J. Clin. Pathol. 56, 699–702. Wattanagoon, Y., Bunnag, D., 2013. Intestinal Fluke Infections. In: Magill, A.J., Ryan, E.T., Hill, D.R., Solomon, T. (Eds.), Hunter’s Tropical Medicine and Emerging Infectious Disease, ninth ed. Saunders Elsevier, China, pp. 884–886, chapter 123. Weinrauch, Y., Zychlinsky, A., 1999. The induction of apoptosis by bacterial pathogens. Annu. Rev. Microbiol. 53, 155–187. Yaguchi, T., Fujikawa, H., Nishizaki, T., 2010. Linoleic acid derivative DCP-LA protects neurons from oxidative stress-induced apoptosis by inhibiting caspase-3/-9 activation. Neurochem. Res. 35, 712–717. Yokogawa, M., Sano, M., 1968. Studies on the intestinal flukes. IV. On the development of the worm in the experimentally infected animals, with metacercariae of Metagonimus yokogawai. Jpn. J. Parasitol. 17, 540. Yu, J.R., Myong, N., Chai, J.Y., 1997. Expression patterns of proliferating cell nuclear antigen in the small intestine of mice infected with Metagonimus yokogawai and Metagonimus Miyata type. Korean J. Parasitol. 35, 239–244.

Insights into regulatory molecules of intestinal epithelial cell turnover during experimental infection by Heterophyes heterophyes.

Heterophyiasis is an intestinal disease that remains endemic in many parts of the world, particularly the Nile Delta of Egypt and Southeast Asia, yet ...
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