15.10. 1975
Specialia
1135
The influence of the i.m. administration of sex hormones in rats on plasma urate levels and urinary excretion of uric acid and allantoin Hormones
No. of animals
Androgeu (10 mg/kg/day)
16
Progesterone (100 mg/kg/day)
13
Estrogen (100 mg/kg/day)
13
Plasma urate level (mg/100 ml)
before after before after before after
2.10 =E 0.81 2.24 4- 0.84 2.43 4- 1.04 1.53 ~2 0.57 ~ 1.85 ~ 0.76 1.87 • 0.93
Urinary excretion Uric acid (mg/day)
Allantoin (mglday)
1.73 J= 1.93 1.82 4- 0.55 1,25 4- 0.21 1,21 4- 0.26 1.99 ~ 0.24 1.97 ! 0.61
72.6 5[2 12.4 75.2 ~ 14.1 58.9 2~ 19.6 53.5 ~= 16.3 70.9 ~= 11.3 73.2 ~2 11.1
~p < 0.05. Plasma urate levels were average values =t- SD of animals. Urinary excretion were shown by average values ~ SD of successioi~ 7 days.
t h a t t h e renal clearance of u r a t e is increased in p r e g n a n c y a n d t h e e n h a n c e d r e n a l e x c r e t i o n of uric acid a n d / o r h y p e r v o l a e m i a of p r e g n a n c y m a y result in t h e h y p o uricemia. H o w e v e r , in e a r l y p r e g n a n c y t h e y could n o t s h o w a n y c h a n g e in r e n a l e x c r e t i o n of uric acid. R e c e n t l y NICHOLLS el: al. ~ e x a m i n e d t h e effect of e s t r o g e n on p l a s m a a n d u r i n a r y uric acid in t r a n s - s e x u a l men, a n d s h o w e d t h a t stilboestrol increases t h e renal e x c r e t i o n of uric acid a n d lowers t h e p l a s m a u r a t e level. T h e y suggested t h a t uricosuric a c t i o n of stilboestrol is a possible cause for age a n d sex differences in p l a s m a uric acid. I n rats, t h e e n d p r o d u c t of p u r i n e m e t a b o l i s m is allant o i n a n d is e x c r e t e d in u r i n e in large a m o u n t s t h o u g h u r i n a r y e x c r e t i o n of uric acid is v e r y little. Therefore, it is difficult to e s t i m a t e t h e renal clearance of uric acid exactly. The p r e s e n t s t u d y s e e m e d to i n d i c a t e t h a t e s t r o g e n h a s no influence on u r a t e p r o d u c t i o n a n d on t h e r e n a l clearance of uric acid. H o w e v e r , it c a n n o t be d e n i e d t h a t e s t r o g e n m a y influence uric acid excretion. W e d e m o n s t r a t e d t h a t p l a s m a u r a t e c o n c e n t r a t i o n and u r i n a r y e x c r e t i o n of uric acid plus allantoin was r e d u c e d b y p r o g e s t e r o n e , a n d t h e r e f o r e it could be a s s u m e d t h a t progesterone diminishes purine synthesis and this effect of p r o g e s t e r o n e would a p p e a r to e x p l a i n t h e sex difference in u r a t e levels in adults. ACH>=SON6 s h o w e d p r e v i o u s l y a significant positive corr e l a t i o n b e t w e e n s e r u m uric acid a n d h a e m o g l o b i n levels in h e a l t h y a d u l t s a n d s u g g e s t e d t h a t t h e sex difference in
u r a t e m e t a b o l i s m m a y be r e l a t e d t o t h e difference in t u r n - o v e r r a t e of blood ceils in b o t h sexes. MIKKELSEN e t al. f o u n d t h a t in male s u b j e c t s t h e p l a s m a u r a t e levels s h o w e d a m a r k e d rise a t t h e p u b e r t y a n d s p e c u l a t e d t h a t a n d r o g e n m a y p l a y an i m p o r t a n t role in raising t h e p l a s m a u r a t e levels. H o w e v e r , we could n o t c o n f i r m t h i s h y p o t h esis in t h e p r e s e n t s t u d y , a n d f u r t h e r e x p e r i m e n t s will h a v e to be carried o u t t o clarify t h e effect of a n d r o g e n on purine metabolism.
Summary. T h e effects of sex h o r m o n e s on p u r i n e m e t a b o l i s m were i n v e s t i g a t e d in rats. N o influence on p u r i n e s y n t h e s i s was s h o w n b y t h e i n j e c t i o n of e s t r o g e n a n d androgen. The p l a s m a u r a t e levels were s i g n i f i c a n t l y lowered f r o m 2.43 • 1.04 mg/100 m l to 1.53 i 0.57 m g / 100 m i b y t h e i n j e c t i o n of p r o g e s t e r o n e . U r i n a r y e x c r e t i o n of uric acid plus a l l a n t o i n was s l i g h t l y r e d u c e d . T h e s e results s u g g e s t e d t h a t p r o g e s t e r o n e m a y influence age a n d sex differences in h u m a n p l a s m a u r a t e levels. Y. NISHIDA, I. AKAOKA a n d T. NISHIZAWA
1)epartment o/Medicine and Physical Therapy, Faculty o] Medicine, University of Tokyo, Bunkyo-ku, Tokyo (Japan 713), 3 June 7975. 5 A. NICIlOLLS,M. L. SNAITIt and J. T. SCOTT, Br. reed. J. 1,449 (1973). 6 R. M. ACHESOSand W. M. O'BRIEX, Lancet 2, 777 (1966).
T h e E f f e c t of H u n g e r o n F r e e F a t t y A c i d a n d C o r t i c o s t e r o n e P l a s m a L e v e l s i n R a t s F a t t y acids p r o v i d e a n e n e r g y source c a p a b l e of r a p i d m o b i l i z a t i o n for n e a r l y all of an o r g a n i s m tissues. T h e y are s t o r e d in t h e adipose tissue as t r i g l y c e r i d e s a n d are set free b y lipolysis. W h e n t h e o r g a n i s m r e q u i r e s m o r e e n e r g y t h a n is available f r o m its diet, or f r o m liver or muscle glycogen, m o b i l i z a t i o n of f a t t y acids is increased 1, 2. The h o r m o n e s of t h e p i t u i t a r y gland a n d t h e a d r e n a l c o r t e x p l a y a n i m p o r t a n t role in t h e m o b i l i z a t i o n of f a t in f a s t i n g a n i m a l s 1,a-~. This i n v e s t i g a t i o n deals w i t h t h e c h a n g e s w i t h time in t h e p l a s m a c o n c e n t r a t i o n s of free f a t t y acids (FFA) a n d c o r t i c o s t e r o n e in t h e case of f a s t i n g r a t s in c o m p a r i s o n w i t h n o r m a l l y fed controls. The d i u r n a l cycles of t h e a n i m a l s were t a k e n i n t o c o n s i d e r a t i o n . Also, t h e effect of h y p o x i a on t h e c o n c e n t r a t i o n c h a n g e s of t h e F F A was investigated.
Material and methods. Male W i s t a r r a t s (160-200 g) were fed a s t a n d a r d d i e t ( R a t b i s c u i t f r o m t h e Tagger Co., Graz) for 4 weeks before t h e s t a r t of testing. The feed c o n t a i n e d 59.5% c a r b o h y d r a t e s , 21.7% p r o t e i n , 2.6% fats, a n d was e n r i c h e d w i t h v i t a m i n s a n d minerals. I n all tests, w a t e r was available t o t h e a n i m a l s ad l i b i t u m . T h e s a m e a n i m a l s were used for all e x p e r i m e n t s . Once a 1 G. F. CAHILL JR., ~V[.G. HERRERA, A. P. MORGAN, J. S. SOELDNER, J. STEINKE, P. L. LEVY, G. A. DxEICHARD JR. and D. lV[. KIv~is,
J. din. Invest. ~5, 1751 (1966). 2 D. S. FREDRICKSONand R. S. GORDON JR., Physiol. Rev. 38, 585 (1958). 3 j. N. FAIN, Endocrinology 71,633 (1962). 4 H..'VI. GOODMANand E. KNOI~IL, Am. J. Physiol. 201, 1 (1961). .5 R. O. Scow and S. S. CHERNICK, Ain. J. Physiol. 270, 1 (1966).
1136
Specialia
w e e k a p p r o x i m a t e l y 1.5 m l blood was t a k e n f r o m t h e vein of t h e t o n g u e , while t h e a n i m a l was u n d e r l i g h t h a l o t h a n e anesthesia% The h y p o x i a e x p e r i m e n t s were cond u c t e d in a c h a m b e r a t a p r e s s u r e of 380 m m Hg, corr e s p o n d i n g t o half a t m o s p h e r i c p r e s s u r e L T h e p l a s m a was d e e p - f r o z e n a t - - 2 5 ~ a n d a n a l y z e d w i t h i n 3 days. The m o d i f i e d D u n c o m b e - m e t h o d a c c o r d i n g t o FAL~OLT e t al. s w a s u s e d for t h e d e t e r m i n a t i o n of F F A . T h e s t a n d a r d c u r v e was c o n s t r u c t e d f r o m a p p r o p r i a t e
1.4
a) "~ 0.8 E
0.6 0.4
b)
0.2 06.00 09[00 12.00 15.00 18.00 21.00 24.00 03.00 06DO Clock tim~ Fig. 1. Concentrations of free fatty acids in the plasma of fed (b) and fasting (a) rats. Each point represents an average value for 5 animals and is plotted together with its standard deviation.
1.0'
0.9 0.8 -g 0.7 0.6 -~ 0.5 0.4 r a--
0,3i 0.2 0.1'
06.00 09.00 12.00 15.00 18.00 2t.00 24.00 05.00 06.00 Clock time Fig. 2. Concentrations of free fatty acids in the plasma of rats during hypoxia. Each point represents an average value for 5 animals and is plotted together with its standard deviation.
400
1
EXPERIENTIA31/10
dilutions of a 0.2 m M p a l m i t i c acid s o l u t i o n t o give a conc e n t r a t i o n r a n g e of 0.01 t o 0.05 ~ M p e r s a m p l e volume, a n d was linear in t h i s range. The s t a n d a r d w h i c h accompanied the determination had a standard deviation
of • 5%. T h e c o r t i c o s t e r o n e c o n t e n t of t h e p l a s m a was d e t e r m i n e d r a d i o i m m u n o l o g i c a l l y using a t e s t kit f r o m t h e Sorin Co., Saluggia, I t a l y , a n d m e a s u r e d w i t h a B e c k m a n L S 230 fluid scintillation c o u n t e r . Results a n d discussion. I t is k n o w n t h a t t h e c o n c e n t r a t i o n s of v a r i o u s p l a s m a c o m p o n e n t s are s u b j e c t to d i u r n a l cycles 9. Therefore, all t e s t s were s t a r t e d a t 06.00 h. W h e n one o b s e r v e s t h e p l a s m a levels of F F A of fed a n i m a l s o v e r a 24-hour period (Figure l b ) , one sees t h a t t h e r e are no g r e a t f l u c t u a t i o n s in t h e curve. Two m a x i m a are seen a t 21.00 a n d 06.00 h a n d 2 m i n i m a a t 15.00 a n d 24.00 h. The c u r v e reflects t h e r e l a t i v e l y large a c t i v i t y c o n n e c t e d w i t h t h e n i g h t l y feeding. F r o m F i g u r e l a one can see t h a t t h e v a l u e s of t h e p l a s m a levels of t h e f a s t i n g a n i m a l s all lie far a b o v e t h o s e for t h e fed animals. The c u r v e shows 2 peaks, one b e t w e e n 12.00 a n d 15.00 h, a n d a n o t h e r a r o u n d m i d n i g h t . T h e m i d n i g h t p e a k reaches a v a l u e of 4 t i m e s t h a t of t h e s t a r t i n g level. A m i n i m u m lies a r o u n d 18.00 h. Similar results were o b t a i n e d b y o t h e r s lo, 11. A simple m e t h o d t o affect a r a p i d m o b i l i z a t i o n of fat, in c o n t r a s t t o t h e g r a d u a l f a t m o b i l i z a t i o n b y h u n g e r , is t h e use of h y p o x i a stress. D u r i n g h y p o x i a t h e c u r v e a l r e a d y r e a c h e s its p e a k value of 4 t i m e s t h e s t a r t i n g level a f t e r 2 h (Figure 2). The c o r t i c o s t e r o n e level of t h e fed a n i m a l s is also s u b j e c t to a d i u r n a l r h y t h m , w i t h a m a x i m u m a t 18.00 h (Figure 3b). The f a s t i n g c u r v e clearly lies h i g h e r t h a n t h a t of t h e fed a n i m a l s a t all points, a n d a l r e a d y reaches its m a x i m u m a t 15.00 h (Figure 3a). T h e m a n y l i p o l y s i s - s t i m u l a t i n g h o r m o n e s can be d i v i d e d into 2 g r o u p s , n a m e l y i n t o r a p i d l y b u t s h o r t a c t i n g h o r m o n e s , a n d slowly b u t long a c t i n g ones. I t can be r e g a r d e d as c e r t a i n t h a t t h e s e 2 groups e x e r t t h e i r effects o v e r s e p a r a t e m e c h a n i s m s . W h i l e t h e r a p i d l y a c t i n g lipolytic h o r m o n e s s t i m u l a t e lipolysis via cyclic a d e n o s i n e m o n o p h o s p h a t e 12, t h e slowly a c t i n g ones affect t h e s y n t h e s i s of R N A a n d p r o t e i n s is. C o r t i c o s t e r o n e belongs in t h e l a t t e r group. A c o m p a r i s o n of t h e plots of t h e curves (Figures l a a n d 3a) s h o w s t h a t t h e cortic o s t e r o n e p e a k occurs before t h e m a x i m u m p l a s m a conc e n t r a t i o n of free f a t t y acids in t i m e . W h i l e t h e increased F F A level a f t e r 12 h of f a s t i n g can be a t t r i b u t e d to an increase in lipolysis d u e t o h o r m o n a l influence, t h e first m a x i m u m a f t e r 6 h of f a s t i n g is p r o b a b l y caused b y a decrease in t h e r e - e s t e r i f i c a t i o n of F F A in t h e f a t cells due to a lack of g l y c e r o l - 3 - p h o s p h a t e . A n increase in lipolysis, therefore, b e g i n s only a f t e r a p p r o x i m a t e l y 12 h,
3oo 6 W. i~/[LEKUSCH,B. PALI~TTA, P. TOFIC und W. TRUPPE, E x p e r i e n t i a
o
28, 870 (1972).
200
? B. PALETTA, Z. Biol. 116, 399 (1971).
oa
100
N o_ i
i
i
06.00 09'.00 12.00 15.00 18.00 21'.00 24.00 0;.00 06'.00 Clock time Fig. 3. Concentrations of corticosterone in the plasma of fed (b) and fasting (a) rats. Each point represents an average value for 8-10 animals and is plotted together with its standard deviation.
s K. FALHOLT, B. LUND and W. FALHOLT,Clin. ehim. Acta 46, 105 (1973). 9 B. E. STATLAND, P. WINKEI. and H. BOKELUND, Clin. Chem. 79, 1374 (1973). 10 W. M. BORTZ and L. A. STE~LX, Biochim. biophys. Aeta 306, 85 (1973). 11 j. D. McGAREY, J. M. MEIER and D. W. FOSTER, J. biol. Chem. 2d8, 270 (1973). 13 R. W. BUTCHERand E. W. SUTHERLAND,J. biok Chem. 237, 1244 (1962). 18 j. N. FAIN, V. P. KOVACEVand R. O. Scow, J. biol. Chem. 240, 3522 (1965).
15.10. 1975
Specialia
w h e n t h e liver a n d muscle glycogen supplies h a v e b e e n exhausted.
Summary. T h e t i m e f u n c t i o n of free f a t t y acids a n d of c o r t i c o s t e r o n e in p l a s m a in fed a n d f a s t e d r a t s was inv e s t i g a t e d . Also a s t u d y of t h e influence of h y p o x i a o n t h e c o n c e n t r a t i o n of p l a s m a free f a t t y acids was c a r r i e d out. W h e r e a s f a s t i n g does n o t seem to s t i m u l a t e lipolysis m a r k e d l y before 12 h, 2 h of h y p o x i a e l e v a t e t h e free
1137
f a t t y acid level to its m a x i m u m . I n t h e f a s t e d c o n d i t i o n , all values are s i g n i f i c a n t l y h i g h e r t h a n c o r t i c o s t e r o n e in t h e fed a n i m a l . W. MLEKUSCH, ~V. TRUPPE, ~V. BEYER u n d B. PALETTA
lnstitut /i~r Medizinische Chemie und -Pregl-Labor der Universitiit Graz, Universitdtsplatz 2, A-8010 Graz (Austria), 76 April 7975.
E v i d e n c e for P u r i n e B i o s y n t h e s i s in H u m a n L e u k o c y t e s P r e v i o u s studies h a v e s h o w n t h a t i s o l a t e d h u m a n l e u k o c y t e s are able to i n c o r p o r a t e 14C-labelled f o r m a t e a n d a d e n i n e into t h e i r nucleic acid b a s e s 1. DIAMOND e t al. 2 o b s e r v e d also a n increased 14C-glycine i n c o r p o r a t i o n i n t o a d e n i n e a n d g u a n i n e of R N A a n d D N A of l e u k o c y t e s f r o m g o u t y p a t i e n t s . H o w e v e r , a d e n i n e a n d g u a n i n e are n o t m e t a b o l i c e n d p r o d u c t s of p u r i n e m e t a b o l i s m a n d t h e i r c o n c e n t r a t i o n c a n n o t give a d e q u a t e i n f o r m a t i o n on t h e b e h a v i o u r of p u r i n e m e t a b o l i s m ; m o r e o v e r , t h e glycine i n c o r p o r a t i o n r a t e d e t e r m i n e d in l e u k o c y t e s in v i t r o does n o t necessarily reflect t h e a c t u a l i m p o r t a n c e of de n o v o p u r i n e b i o s y n t h e s i s . R e c e n t l y , REaM a f o u n d t h a t B u r k i t t l y m p h o m a a n d h u m a n spleen cells could s y n t h e t i z e t h e first a n d second i n t e r m e d i a t e c o m p o u n d s of t h e p u r i n e b i o s y n t h e t i c p a t h w a y , a n d VV'OOD a n d S~2EOMILLER4 a s s a y e d a n d d e t e r m i n e d some p r o p e r t i e s of 5 - p h o s p h o r i bosyl-l-pyrophosphate amidotransferase, the enzyme t h a t c a t a l y z e s t h e first step i n v o l v e d in de n o v o p u r i n e b i o s y n t h e s i s , in h u m a n l y m p h o b l a s t s m a i n t a i n e d in tissue culture. T h e p u r p o s e of t h e p r e s e n t s t u d y was to investigate whether human leukocytes and lymphocytes are c a p a b l e of de n o v o p u r i n e b i o s y n t h e s i s . ~Ve a s s a y e d t h e a c t i v i t y of 5 - p h o s p h o r i b o s y l - l - p y r o p h o s p h a t e a m i d o t r a n s f e r a s e in l e u k o c y t e s a n d l y m p h o c y t e s isolated b y t h e u s u a l p r o c e d u r e s from 11 h e a l t h y v o l u n t e e r s u b j e c t s a c c o r d i n g to t h e m e t h o d of WYNOAARDEN a n d ASHTON a. T h e assays were p e r f o r m e d on l e u k o c y t e s a n d l y m p h o c y t e s e x t r a c t s dialyzed 24 h a g a i n s t distilled w a t e r in order to r e m o v e h e p a r i n a n d o t h e r molecules i n t e r f e r i n g or i n h i b i t i n g t h e e n z y m e . E n z y m e a c t i v i t y was expressed as units/10" cells, w h e r e one u n i t of e n z y m e a c t i v i t y is t h e a m o u n t t h a t p r o d u c e s a n increase of 0.001 a b s o r b a n c e u n i t / m i n a t 363 n m a t p H 8 a n d r o o m t e m p e r a t u r e (25 ~ E n z y m e a c t i v i t y h a s b e e n d e t e c t e d in b o t h k i n d s of cells e x a m i n e d . T h e m e a n v a l u e s r e s u l t e d 1.34 ~ 0.58 units/106 cells in l e u k o c y t e s a n d 0.86 ~ 0.66 units/106 cells in l y m p h o c y t e s (Table). P r e f o r m e d p u r i n e s c o n v e r t e d b y cellular m e t a b o l i s m to r i b o n u c l e o t i d e s h a v e b e e n c o n s i d e r e d t h e o n l y source of p u r i n e n u c l e o t i d e s for m a n y h u m a n cellsa,% Most of t h e conclusions on t h i s d e p e n d e n c e are, h o w e v e r , b a s e d o n i n d i r e c t or n e g a t i v e evidence. S t u d y of t h e e a r l y steps of de n o v o p u r i n e b i o s y n t h e s i s in h u m a n cells, such 5-Phosphoribosyl-l-pyrophosphate amidotransferase activity in leukocytes and lymphoeytes of healthy human subjects
as fibroblasts, l e u k o c y t e s a n d platelets, h a s b e e n l i m i t e d to e v a l u a t i n g t h e a c c u m u l a t i o n of f o r m y l g l y c i n a m i d e r i b o n u c l e o t i d e 3,6 or o b s e r v i n g t h e failure to i n c o r p o r a t e glycine or f o r m a t e i n t o p u r i n e n u c l e o t i d e s in v i t r o 6, while t h e p r e s e n c e a n d t h e p r o p e r t i e s of 5 - p h o s p h o r i b o s y l - 1 pyrophosphate amidotransferase have not been extensively e x a m i n e d 4. H o w e v e r , t h e i n t e r p r e t a t i o n of isotopei n c o r p o r a t i o n studies i n v o l v i n g p u r i n e s y n t h e s i s r e a c t i o n s requires some care c o n c e r n i n g t h e c o n c e n t r a t i o n of precursors a n d i n t e r m e d i a t e c o m p o u n d s 1, 6, a n d t h e p o s s i b i l i t y t h a t some s u b s t r a t e n e c e s s a r y for de n o v o s y n t h e s i s m a y b e e n lacking% O u r results p r o v i d e e v i d e n d e t h a t h u m a n l e u k o c y t e s a n d l y m p h o c y t e s poss2ss t h e e n z y m e a c t i v i t y t h a t c a t a l y z e s t h e f o r m a t i o n of t h e first i n t e r m e d i a t e c o m p o u n d of t h e p u r i n e b i o s y n t h e t i c p a t h w a y . T h e f i n d i n g of t h e p r e s e n c e of 5 - p h o s p h o r i b o s y l - l - p y r o p h o s p h a t e a m i d o t r a n s f e r a s e in h u m a n l e u k o c y t e s a n d l y m p h o c y t e s suggests, therefore, t h a t t h e s e cells h a v e t h e c a p a c i t y for de n o v o p u r i n e b i o s y n t h e s i s a n d are n o t solely d e p e n d e n t on t h e liver for t h e i r s u p p l y of purines. De n o v o p u r i n e s y n t h e s i s m a y t h e r e f o r e be more w i d e s p r e a d than previously reported, and human leukocytes and l y m p h o c y t c s m a y be a n i m p o r t a n t e x t r a h e p a t i c site for de n o v e p u r i n e b i o s y n t h e s i s in m a n . T h e s t u d y of t h e p u r i n e b i o s y n t h e t i c p a t h w a y in h u m a n l e u k o c y t e s a n d l y m p h o c y t e s seems of p a r t i c u l a r i n t e r e s t since i t could p r o v i d e a v a l u a b l e e x p e r i m e n t a l m o d e l to i n v e s t i g a t e t h e factors i n v o l v e d in t h e r e g u l a t i o n of de n o v o p u r i n e bios y n t h e s i s a n d clarify p u r i n e m e t a b o l i s m a l t e r a t i o n s in p a t i e n t s w i t h h y p e r u r i c e m i a , gout, or l e u k e m i a a n d o t h e r m y e l o p r o l i f e r a t i v e diseases.
Summary. H u m a n l e u k o c y t e s a n d l y m p h o c y t e s h a v e s h o w n to b e e q u i p p 3 d w i t h 5 - p h o s p h o r i b o s y l - l - p y r o p h o s phate amidotransrease, the enzyme which catalyzes the s y n t h e s i s of t h e first i n t e r m e d i a t e of t h e p u r i n e p a t h w a y , t h u s p r o v i d i n g e v i d e n c e t h a t t h e s e cells h a v e t h e c a p a c i t y for de n o v o p u r i n e b i o s y n t h e s i s . R. MARCOLONGO, G. POMPUCCI a n d V. MICHELI
Cattedra di Reumatologia and Institute o/ Biochemistry, University o/ Siena, Via Ponizetti 7, Siena (italy), 26 May 7975. 1 j. L. SCOTT, J. clin. hlvest. 41, 67 (1962). 2 }{. S. DIAMOND, M. FRIEDLANI), D. HOLBERSTAM a n d 1). KAPLAN,
Cell type
Enzyme activity (units/l 0~cells)
Leukocytes Lymphocytes
1.34 -c 0.58 0.86 -c 0.66
Ann. rheum. Dis. 28, 27.5 (1969). 3 G. H. REEM, J. clin. Invest. 57, 1058 (1972). 4 A. W. WOOD and J. E. SEEGMILLER, J. biol. Chem. 248, 138 (1973). J. B. WYNOAAm)EN and D. M. ASHTDN, J. biol. Chem. 234, 1492 (1959). A. \V. MURRAY, Ann. Rev. lgioehem. 40, 811 (1971).
Specialia
1135
The influence of the i.m. administration of sex hormones in rats on plasma urate levels and urinary excretion of uric acid and allantoin Hormones
No. of animals
Androgeu (10 mg/kg/day)
16
Progesterone (100 mg/kg/day)
13
Estrogen (100 mg/kg/day)
13
Plasma urate level (mg/100 ml)
before after before after before after
2.10 =E 0.81 2.24 4- 0.84 2.43 4- 1.04 1.53 ~2 0.57 ~ 1.85 ~ 0.76 1.87 • 0.93
Urinary excretion Uric acid (mg/day)
Allantoin (mglday)
1.73 J= 1.93 1.82 4- 0.55 1,25 4- 0.21 1,21 4- 0.26 1.99 ~ 0.24 1.97 ! 0.61
72.6 5[2 12.4 75.2 ~ 14.1 58.9 2~ 19.6 53.5 ~= 16.3 70.9 ~= 11.3 73.2 ~2 11.1
~p < 0.05. Plasma urate levels were average values =t- SD of animals. Urinary excretion were shown by average values ~ SD of successioi~ 7 days.
t h a t t h e renal clearance of u r a t e is increased in p r e g n a n c y a n d t h e e n h a n c e d r e n a l e x c r e t i o n of uric acid a n d / o r h y p e r v o l a e m i a of p r e g n a n c y m a y result in t h e h y p o uricemia. H o w e v e r , in e a r l y p r e g n a n c y t h e y could n o t s h o w a n y c h a n g e in r e n a l e x c r e t i o n of uric acid. R e c e n t l y NICHOLLS el: al. ~ e x a m i n e d t h e effect of e s t r o g e n on p l a s m a a n d u r i n a r y uric acid in t r a n s - s e x u a l men, a n d s h o w e d t h a t stilboestrol increases t h e renal e x c r e t i o n of uric acid a n d lowers t h e p l a s m a u r a t e level. T h e y suggested t h a t uricosuric a c t i o n of stilboestrol is a possible cause for age a n d sex differences in p l a s m a uric acid. I n rats, t h e e n d p r o d u c t of p u r i n e m e t a b o l i s m is allant o i n a n d is e x c r e t e d in u r i n e in large a m o u n t s t h o u g h u r i n a r y e x c r e t i o n of uric acid is v e r y little. Therefore, it is difficult to e s t i m a t e t h e renal clearance of uric acid exactly. The p r e s e n t s t u d y s e e m e d to i n d i c a t e t h a t e s t r o g e n h a s no influence on u r a t e p r o d u c t i o n a n d on t h e r e n a l clearance of uric acid. H o w e v e r , it c a n n o t be d e n i e d t h a t e s t r o g e n m a y influence uric acid excretion. W e d e m o n s t r a t e d t h a t p l a s m a u r a t e c o n c e n t r a t i o n and u r i n a r y e x c r e t i o n of uric acid plus allantoin was r e d u c e d b y p r o g e s t e r o n e , a n d t h e r e f o r e it could be a s s u m e d t h a t progesterone diminishes purine synthesis and this effect of p r o g e s t e r o n e would a p p e a r to e x p l a i n t h e sex difference in u r a t e levels in adults. ACH>=SON6 s h o w e d p r e v i o u s l y a significant positive corr e l a t i o n b e t w e e n s e r u m uric acid a n d h a e m o g l o b i n levels in h e a l t h y a d u l t s a n d s u g g e s t e d t h a t t h e sex difference in
u r a t e m e t a b o l i s m m a y be r e l a t e d t o t h e difference in t u r n - o v e r r a t e of blood ceils in b o t h sexes. MIKKELSEN e t al. f o u n d t h a t in male s u b j e c t s t h e p l a s m a u r a t e levels s h o w e d a m a r k e d rise a t t h e p u b e r t y a n d s p e c u l a t e d t h a t a n d r o g e n m a y p l a y an i m p o r t a n t role in raising t h e p l a s m a u r a t e levels. H o w e v e r , we could n o t c o n f i r m t h i s h y p o t h esis in t h e p r e s e n t s t u d y , a n d f u r t h e r e x p e r i m e n t s will h a v e to be carried o u t t o clarify t h e effect of a n d r o g e n on purine metabolism.
Summary. T h e effects of sex h o r m o n e s on p u r i n e m e t a b o l i s m were i n v e s t i g a t e d in rats. N o influence on p u r i n e s y n t h e s i s was s h o w n b y t h e i n j e c t i o n of e s t r o g e n a n d androgen. The p l a s m a u r a t e levels were s i g n i f i c a n t l y lowered f r o m 2.43 • 1.04 mg/100 m l to 1.53 i 0.57 m g / 100 m i b y t h e i n j e c t i o n of p r o g e s t e r o n e . U r i n a r y e x c r e t i o n of uric acid plus a l l a n t o i n was s l i g h t l y r e d u c e d . T h e s e results s u g g e s t e d t h a t p r o g e s t e r o n e m a y influence age a n d sex differences in h u m a n p l a s m a u r a t e levels. Y. NISHIDA, I. AKAOKA a n d T. NISHIZAWA
1)epartment o/Medicine and Physical Therapy, Faculty o] Medicine, University of Tokyo, Bunkyo-ku, Tokyo (Japan 713), 3 June 7975. 5 A. NICIlOLLS,M. L. SNAITIt and J. T. SCOTT, Br. reed. J. 1,449 (1973). 6 R. M. ACHESOSand W. M. O'BRIEX, Lancet 2, 777 (1966).
T h e E f f e c t of H u n g e r o n F r e e F a t t y A c i d a n d C o r t i c o s t e r o n e P l a s m a L e v e l s i n R a t s F a t t y acids p r o v i d e a n e n e r g y source c a p a b l e of r a p i d m o b i l i z a t i o n for n e a r l y all of an o r g a n i s m tissues. T h e y are s t o r e d in t h e adipose tissue as t r i g l y c e r i d e s a n d are set free b y lipolysis. W h e n t h e o r g a n i s m r e q u i r e s m o r e e n e r g y t h a n is available f r o m its diet, or f r o m liver or muscle glycogen, m o b i l i z a t i o n of f a t t y acids is increased 1, 2. The h o r m o n e s of t h e p i t u i t a r y gland a n d t h e a d r e n a l c o r t e x p l a y a n i m p o r t a n t role in t h e m o b i l i z a t i o n of f a t in f a s t i n g a n i m a l s 1,a-~. This i n v e s t i g a t i o n deals w i t h t h e c h a n g e s w i t h time in t h e p l a s m a c o n c e n t r a t i o n s of free f a t t y acids (FFA) a n d c o r t i c o s t e r o n e in t h e case of f a s t i n g r a t s in c o m p a r i s o n w i t h n o r m a l l y fed controls. The d i u r n a l cycles of t h e a n i m a l s were t a k e n i n t o c o n s i d e r a t i o n . Also, t h e effect of h y p o x i a on t h e c o n c e n t r a t i o n c h a n g e s of t h e F F A was investigated.
Material and methods. Male W i s t a r r a t s (160-200 g) were fed a s t a n d a r d d i e t ( R a t b i s c u i t f r o m t h e Tagger Co., Graz) for 4 weeks before t h e s t a r t of testing. The feed c o n t a i n e d 59.5% c a r b o h y d r a t e s , 21.7% p r o t e i n , 2.6% fats, a n d was e n r i c h e d w i t h v i t a m i n s a n d minerals. I n all tests, w a t e r was available t o t h e a n i m a l s ad l i b i t u m . T h e s a m e a n i m a l s were used for all e x p e r i m e n t s . Once a 1 G. F. CAHILL JR., ~V[.G. HERRERA, A. P. MORGAN, J. S. SOELDNER, J. STEINKE, P. L. LEVY, G. A. DxEICHARD JR. and D. lV[. KIv~is,
J. din. Invest. ~5, 1751 (1966). 2 D. S. FREDRICKSONand R. S. GORDON JR., Physiol. Rev. 38, 585 (1958). 3 j. N. FAIN, Endocrinology 71,633 (1962). 4 H..'VI. GOODMANand E. KNOI~IL, Am. J. Physiol. 201, 1 (1961). .5 R. O. Scow and S. S. CHERNICK, Ain. J. Physiol. 270, 1 (1966).
1136
Specialia
w e e k a p p r o x i m a t e l y 1.5 m l blood was t a k e n f r o m t h e vein of t h e t o n g u e , while t h e a n i m a l was u n d e r l i g h t h a l o t h a n e anesthesia% The h y p o x i a e x p e r i m e n t s were cond u c t e d in a c h a m b e r a t a p r e s s u r e of 380 m m Hg, corr e s p o n d i n g t o half a t m o s p h e r i c p r e s s u r e L T h e p l a s m a was d e e p - f r o z e n a t - - 2 5 ~ a n d a n a l y z e d w i t h i n 3 days. The m o d i f i e d D u n c o m b e - m e t h o d a c c o r d i n g t o FAL~OLT e t al. s w a s u s e d for t h e d e t e r m i n a t i o n of F F A . T h e s t a n d a r d c u r v e was c o n s t r u c t e d f r o m a p p r o p r i a t e
1.4
a) "~ 0.8 E
0.6 0.4
b)
0.2 06.00 09[00 12.00 15.00 18.00 21.00 24.00 03.00 06DO Clock tim~ Fig. 1. Concentrations of free fatty acids in the plasma of fed (b) and fasting (a) rats. Each point represents an average value for 5 animals and is plotted together with its standard deviation.
1.0'
0.9 0.8 -g 0.7 0.6 -~ 0.5 0.4 r a--
0,3i 0.2 0.1'
06.00 09.00 12.00 15.00 18.00 2t.00 24.00 05.00 06.00 Clock time Fig. 2. Concentrations of free fatty acids in the plasma of rats during hypoxia. Each point represents an average value for 5 animals and is plotted together with its standard deviation.
400
1
EXPERIENTIA31/10
dilutions of a 0.2 m M p a l m i t i c acid s o l u t i o n t o give a conc e n t r a t i o n r a n g e of 0.01 t o 0.05 ~ M p e r s a m p l e volume, a n d was linear in t h i s range. The s t a n d a r d w h i c h accompanied the determination had a standard deviation
of • 5%. T h e c o r t i c o s t e r o n e c o n t e n t of t h e p l a s m a was d e t e r m i n e d r a d i o i m m u n o l o g i c a l l y using a t e s t kit f r o m t h e Sorin Co., Saluggia, I t a l y , a n d m e a s u r e d w i t h a B e c k m a n L S 230 fluid scintillation c o u n t e r . Results a n d discussion. I t is k n o w n t h a t t h e c o n c e n t r a t i o n s of v a r i o u s p l a s m a c o m p o n e n t s are s u b j e c t to d i u r n a l cycles 9. Therefore, all t e s t s were s t a r t e d a t 06.00 h. W h e n one o b s e r v e s t h e p l a s m a levels of F F A of fed a n i m a l s o v e r a 24-hour period (Figure l b ) , one sees t h a t t h e r e are no g r e a t f l u c t u a t i o n s in t h e curve. Two m a x i m a are seen a t 21.00 a n d 06.00 h a n d 2 m i n i m a a t 15.00 a n d 24.00 h. The c u r v e reflects t h e r e l a t i v e l y large a c t i v i t y c o n n e c t e d w i t h t h e n i g h t l y feeding. F r o m F i g u r e l a one can see t h a t t h e v a l u e s of t h e p l a s m a levels of t h e f a s t i n g a n i m a l s all lie far a b o v e t h o s e for t h e fed animals. The c u r v e shows 2 peaks, one b e t w e e n 12.00 a n d 15.00 h, a n d a n o t h e r a r o u n d m i d n i g h t . T h e m i d n i g h t p e a k reaches a v a l u e of 4 t i m e s t h a t of t h e s t a r t i n g level. A m i n i m u m lies a r o u n d 18.00 h. Similar results were o b t a i n e d b y o t h e r s lo, 11. A simple m e t h o d t o affect a r a p i d m o b i l i z a t i o n of fat, in c o n t r a s t t o t h e g r a d u a l f a t m o b i l i z a t i o n b y h u n g e r , is t h e use of h y p o x i a stress. D u r i n g h y p o x i a t h e c u r v e a l r e a d y r e a c h e s its p e a k value of 4 t i m e s t h e s t a r t i n g level a f t e r 2 h (Figure 2). The c o r t i c o s t e r o n e level of t h e fed a n i m a l s is also s u b j e c t to a d i u r n a l r h y t h m , w i t h a m a x i m u m a t 18.00 h (Figure 3b). The f a s t i n g c u r v e clearly lies h i g h e r t h a n t h a t of t h e fed a n i m a l s a t all points, a n d a l r e a d y reaches its m a x i m u m a t 15.00 h (Figure 3a). T h e m a n y l i p o l y s i s - s t i m u l a t i n g h o r m o n e s can be d i v i d e d into 2 g r o u p s , n a m e l y i n t o r a p i d l y b u t s h o r t a c t i n g h o r m o n e s , a n d slowly b u t long a c t i n g ones. I t can be r e g a r d e d as c e r t a i n t h a t t h e s e 2 groups e x e r t t h e i r effects o v e r s e p a r a t e m e c h a n i s m s . W h i l e t h e r a p i d l y a c t i n g lipolytic h o r m o n e s s t i m u l a t e lipolysis via cyclic a d e n o s i n e m o n o p h o s p h a t e 12, t h e slowly a c t i n g ones affect t h e s y n t h e s i s of R N A a n d p r o t e i n s is. C o r t i c o s t e r o n e belongs in t h e l a t t e r group. A c o m p a r i s o n of t h e plots of t h e curves (Figures l a a n d 3a) s h o w s t h a t t h e cortic o s t e r o n e p e a k occurs before t h e m a x i m u m p l a s m a conc e n t r a t i o n of free f a t t y acids in t i m e . W h i l e t h e increased F F A level a f t e r 12 h of f a s t i n g can be a t t r i b u t e d to an increase in lipolysis d u e t o h o r m o n a l influence, t h e first m a x i m u m a f t e r 6 h of f a s t i n g is p r o b a b l y caused b y a decrease in t h e r e - e s t e r i f i c a t i o n of F F A in t h e f a t cells due to a lack of g l y c e r o l - 3 - p h o s p h a t e . A n increase in lipolysis, therefore, b e g i n s only a f t e r a p p r o x i m a t e l y 12 h,
3oo 6 W. i~/[LEKUSCH,B. PALI~TTA, P. TOFIC und W. TRUPPE, E x p e r i e n t i a
o
28, 870 (1972).
200
? B. PALETTA, Z. Biol. 116, 399 (1971).
oa
100
N o_ i
i
i
06.00 09'.00 12.00 15.00 18.00 21'.00 24.00 0;.00 06'.00 Clock time Fig. 3. Concentrations of corticosterone in the plasma of fed (b) and fasting (a) rats. Each point represents an average value for 8-10 animals and is plotted together with its standard deviation.
s K. FALHOLT, B. LUND and W. FALHOLT,Clin. ehim. Acta 46, 105 (1973). 9 B. E. STATLAND, P. WINKEI. and H. BOKELUND, Clin. Chem. 79, 1374 (1973). 10 W. M. BORTZ and L. A. STE~LX, Biochim. biophys. Aeta 306, 85 (1973). 11 j. D. McGAREY, J. M. MEIER and D. W. FOSTER, J. biol. Chem. 2d8, 270 (1973). 13 R. W. BUTCHERand E. W. SUTHERLAND,J. biok Chem. 237, 1244 (1962). 18 j. N. FAIN, V. P. KOVACEVand R. O. Scow, J. biol. Chem. 240, 3522 (1965).
15.10. 1975
Specialia
w h e n t h e liver a n d muscle glycogen supplies h a v e b e e n exhausted.
Summary. T h e t i m e f u n c t i o n of free f a t t y acids a n d of c o r t i c o s t e r o n e in p l a s m a in fed a n d f a s t e d r a t s was inv e s t i g a t e d . Also a s t u d y of t h e influence of h y p o x i a o n t h e c o n c e n t r a t i o n of p l a s m a free f a t t y acids was c a r r i e d out. W h e r e a s f a s t i n g does n o t seem to s t i m u l a t e lipolysis m a r k e d l y before 12 h, 2 h of h y p o x i a e l e v a t e t h e free
1137
f a t t y acid level to its m a x i m u m . I n t h e f a s t e d c o n d i t i o n , all values are s i g n i f i c a n t l y h i g h e r t h a n c o r t i c o s t e r o n e in t h e fed a n i m a l . W. MLEKUSCH, ~V. TRUPPE, ~V. BEYER u n d B. PALETTA
lnstitut /i~r Medizinische Chemie und -Pregl-Labor der Universitiit Graz, Universitdtsplatz 2, A-8010 Graz (Austria), 76 April 7975.
E v i d e n c e for P u r i n e B i o s y n t h e s i s in H u m a n L e u k o c y t e s P r e v i o u s studies h a v e s h o w n t h a t i s o l a t e d h u m a n l e u k o c y t e s are able to i n c o r p o r a t e 14C-labelled f o r m a t e a n d a d e n i n e into t h e i r nucleic acid b a s e s 1. DIAMOND e t al. 2 o b s e r v e d also a n increased 14C-glycine i n c o r p o r a t i o n i n t o a d e n i n e a n d g u a n i n e of R N A a n d D N A of l e u k o c y t e s f r o m g o u t y p a t i e n t s . H o w e v e r , a d e n i n e a n d g u a n i n e are n o t m e t a b o l i c e n d p r o d u c t s of p u r i n e m e t a b o l i s m a n d t h e i r c o n c e n t r a t i o n c a n n o t give a d e q u a t e i n f o r m a t i o n on t h e b e h a v i o u r of p u r i n e m e t a b o l i s m ; m o r e o v e r , t h e glycine i n c o r p o r a t i o n r a t e d e t e r m i n e d in l e u k o c y t e s in v i t r o does n o t necessarily reflect t h e a c t u a l i m p o r t a n c e of de n o v o p u r i n e b i o s y n t h e s i s . R e c e n t l y , REaM a f o u n d t h a t B u r k i t t l y m p h o m a a n d h u m a n spleen cells could s y n t h e t i z e t h e first a n d second i n t e r m e d i a t e c o m p o u n d s of t h e p u r i n e b i o s y n t h e t i c p a t h w a y , a n d VV'OOD a n d S~2EOMILLER4 a s s a y e d a n d d e t e r m i n e d some p r o p e r t i e s of 5 - p h o s p h o r i bosyl-l-pyrophosphate amidotransferase, the enzyme t h a t c a t a l y z e s t h e first step i n v o l v e d in de n o v o p u r i n e b i o s y n t h e s i s , in h u m a n l y m p h o b l a s t s m a i n t a i n e d in tissue culture. T h e p u r p o s e of t h e p r e s e n t s t u d y was to investigate whether human leukocytes and lymphocytes are c a p a b l e of de n o v o p u r i n e b i o s y n t h e s i s . ~Ve a s s a y e d t h e a c t i v i t y of 5 - p h o s p h o r i b o s y l - l - p y r o p h o s p h a t e a m i d o t r a n s f e r a s e in l e u k o c y t e s a n d l y m p h o c y t e s isolated b y t h e u s u a l p r o c e d u r e s from 11 h e a l t h y v o l u n t e e r s u b j e c t s a c c o r d i n g to t h e m e t h o d of WYNOAARDEN a n d ASHTON a. T h e assays were p e r f o r m e d on l e u k o c y t e s a n d l y m p h o c y t e s e x t r a c t s dialyzed 24 h a g a i n s t distilled w a t e r in order to r e m o v e h e p a r i n a n d o t h e r molecules i n t e r f e r i n g or i n h i b i t i n g t h e e n z y m e . E n z y m e a c t i v i t y was expressed as units/10" cells, w h e r e one u n i t of e n z y m e a c t i v i t y is t h e a m o u n t t h a t p r o d u c e s a n increase of 0.001 a b s o r b a n c e u n i t / m i n a t 363 n m a t p H 8 a n d r o o m t e m p e r a t u r e (25 ~ E n z y m e a c t i v i t y h a s b e e n d e t e c t e d in b o t h k i n d s of cells e x a m i n e d . T h e m e a n v a l u e s r e s u l t e d 1.34 ~ 0.58 units/106 cells in l e u k o c y t e s a n d 0.86 ~ 0.66 units/106 cells in l y m p h o c y t e s (Table). P r e f o r m e d p u r i n e s c o n v e r t e d b y cellular m e t a b o l i s m to r i b o n u c l e o t i d e s h a v e b e e n c o n s i d e r e d t h e o n l y source of p u r i n e n u c l e o t i d e s for m a n y h u m a n cellsa,% Most of t h e conclusions on t h i s d e p e n d e n c e are, h o w e v e r , b a s e d o n i n d i r e c t or n e g a t i v e evidence. S t u d y of t h e e a r l y steps of de n o v o p u r i n e b i o s y n t h e s i s in h u m a n cells, such 5-Phosphoribosyl-l-pyrophosphate amidotransferase activity in leukocytes and lymphoeytes of healthy human subjects
as fibroblasts, l e u k o c y t e s a n d platelets, h a s b e e n l i m i t e d to e v a l u a t i n g t h e a c c u m u l a t i o n of f o r m y l g l y c i n a m i d e r i b o n u c l e o t i d e 3,6 or o b s e r v i n g t h e failure to i n c o r p o r a t e glycine or f o r m a t e i n t o p u r i n e n u c l e o t i d e s in v i t r o 6, while t h e p r e s e n c e a n d t h e p r o p e r t i e s of 5 - p h o s p h o r i b o s y l - 1 pyrophosphate amidotransferase have not been extensively e x a m i n e d 4. H o w e v e r , t h e i n t e r p r e t a t i o n of isotopei n c o r p o r a t i o n studies i n v o l v i n g p u r i n e s y n t h e s i s r e a c t i o n s requires some care c o n c e r n i n g t h e c o n c e n t r a t i o n of precursors a n d i n t e r m e d i a t e c o m p o u n d s 1, 6, a n d t h e p o s s i b i l i t y t h a t some s u b s t r a t e n e c e s s a r y for de n o v o s y n t h e s i s m a y b e e n lacking% O u r results p r o v i d e e v i d e n d e t h a t h u m a n l e u k o c y t e s a n d l y m p h o c y t e s poss2ss t h e e n z y m e a c t i v i t y t h a t c a t a l y z e s t h e f o r m a t i o n of t h e first i n t e r m e d i a t e c o m p o u n d of t h e p u r i n e b i o s y n t h e t i c p a t h w a y . T h e f i n d i n g of t h e p r e s e n c e of 5 - p h o s p h o r i b o s y l - l - p y r o p h o s p h a t e a m i d o t r a n s f e r a s e in h u m a n l e u k o c y t e s a n d l y m p h o c y t e s suggests, therefore, t h a t t h e s e cells h a v e t h e c a p a c i t y for de n o v o p u r i n e b i o s y n t h e s i s a n d are n o t solely d e p e n d e n t on t h e liver for t h e i r s u p p l y of purines. De n o v o p u r i n e s y n t h e s i s m a y t h e r e f o r e be more w i d e s p r e a d than previously reported, and human leukocytes and l y m p h o c y t c s m a y be a n i m p o r t a n t e x t r a h e p a t i c site for de n o v e p u r i n e b i o s y n t h e s i s in m a n . T h e s t u d y of t h e p u r i n e b i o s y n t h e t i c p a t h w a y in h u m a n l e u k o c y t e s a n d l y m p h o c y t e s seems of p a r t i c u l a r i n t e r e s t since i t could p r o v i d e a v a l u a b l e e x p e r i m e n t a l m o d e l to i n v e s t i g a t e t h e factors i n v o l v e d in t h e r e g u l a t i o n of de n o v o p u r i n e bios y n t h e s i s a n d clarify p u r i n e m e t a b o l i s m a l t e r a t i o n s in p a t i e n t s w i t h h y p e r u r i c e m i a , gout, or l e u k e m i a a n d o t h e r m y e l o p r o l i f e r a t i v e diseases.
Summary. H u m a n l e u k o c y t e s a n d l y m p h o c y t e s h a v e s h o w n to b e e q u i p p 3 d w i t h 5 - p h o s p h o r i b o s y l - l - p y r o p h o s phate amidotransrease, the enzyme which catalyzes the s y n t h e s i s of t h e first i n t e r m e d i a t e of t h e p u r i n e p a t h w a y , t h u s p r o v i d i n g e v i d e n c e t h a t t h e s e cells h a v e t h e c a p a c i t y for de n o v o p u r i n e b i o s y n t h e s i s . R. MARCOLONGO, G. POMPUCCI a n d V. MICHELI
Cattedra di Reumatologia and Institute o/ Biochemistry, University o/ Siena, Via Ponizetti 7, Siena (italy), 26 May 7975. 1 j. L. SCOTT, J. clin. hlvest. 41, 67 (1962). 2 }{. S. DIAMOND, M. FRIEDLANI), D. HOLBERSTAM a n d 1). KAPLAN,
Cell type
Enzyme activity (units/l 0~cells)
Leukocytes Lymphocytes
1.34 -c 0.58 0.86 -c 0.66
Ann. rheum. Dis. 28, 27.5 (1969). 3 G. H. REEM, J. clin. Invest. 57, 1058 (1972). 4 A. W. WOOD and J. E. SEEGMILLER, J. biol. Chem. 248, 138 (1973). J. B. WYNOAAm)EN and D. M. ASHTDN, J. biol. Chem. 234, 1492 (1959). A. \V. MURRAY, Ann. Rev. lgioehem. 40, 811 (1971).
