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Inhibition of helper T cell function and expression of “Ia” by histamine induced suppressor factor
562
Annual A A C H T Meeung, 1981
SPECIES CROSS- REACTIVE I a DETERMINANTS. DETECTION BY MONOCLONALANTIBODY TO HLT)La~ I s . JUDY FALK AND HICHELLE LETARTE, TORONTO LW.S,~E~ HOSPITAL AND HOSPITAL FOR S1CK CHILDREN, TORONTO, CANADA. A.TH a n t i - A . T L s e r m l , d i r e c t e d a t t h e I a a n t l g e n s o f murine h a p l o t y p e k . r e a c t s s t r o n g l y v i t h nen-polymorphic d e t e r m i n a n t s on a!1 huemn B c e l l s and B - C L L c e l l s t e s t e d . I a J and B p o l y p e p t l d e c h a i n s can be Immunoprecipltated from c e l l e x t r a c t s by t h e murine I-E/C s u b r e g i o n , ge have nov d e m o n s t r a t e d ~hat A.TH s n t l - A . T L s e r u m r e a c t s w l t h ~heep, p l g and r a t c e l l s a s ~ e l l a s v l t h human c e l l s . I n h i b i t i o n s ~ u g i e s , u s i n g t h e c e l l u l a r radlolmmunoassay, have d e m o n s t r a t e d t h a t s h e e p , p l g and r a t c e l l s c o u l d a b s o r b t h e b i n d i n g o f ~he A . T H a n t i - A . T L a e r u m t o htman I s . Thus t h i s " a l l o s e r u m '~ can a l s o r e c o g n i z e and I s d e t e r m i n a n t ( s ) common t o s e v e r a l s p e c i e s . A monoclonal a n t i b o d y , 21w4, r s l s e d b} I m u n l z l n g A.THmlce w i t h p u r i f i e d human l a g l y c o p r o t e i n s , d e t e c t e d nonpolymorphlc h e , a n l a d e t e t m l n a n t s I n t h e c e l l u l a r radiole~aunoassay; 21w4 a n t i b o d y a l s o r e a c t e d w i t h p i g , sheep and some mouse s t r a i n s . The murine s p e c i f i c i t y r e c o g n i z e d by t b i s monoclonal a n t i b o d y t o human I a m a p p o d t o t h e I-E/C s u b r e g i o n . The murine r e a c t i v i t y o f 21w4 c o u l d be a b s o r b e d by human, s h e e p and p i g c e l l s . T h e r e f o r e Immunization o f mtee of h a p l o t y p e s ( l a c k i n g I - E / C gene p r o d u c t s ) u i t h e l t h e r m u r l n e o r human l a m o l e c u l e s , e l i c i t s a n t i b o d i e s recognlzingamlno a c l d sequences conserve.l t h r o u g h o u t e v o l u t i o n , lqonoclonal a n t l b o d l ~ s such a s 21w4 a r e t h u s n o t o n l y v a l u a b l e f o r t h e puri~leatlonof human I s m o l e c u l e s b u t a l s o f o r t h e c h a r a c t e r l z a t l o n and p u r i f i c a t i o n o f I s m o l e c u l e s from s e v e r a l s p e c i e s .
THE A3,B7 HAPLOTYPE. DECREASEDNATURALKILLER (NK) CELL ACTIVITY IN MULTIPLE SCLEROSIS, H.R._Garovoyo $. Hau~er~ K. AulL. C. Connors~ H.L. We]net. Depts. of Medicine and Neurology, Brigham and Women's Hospttalo Boston, Has-s. Several 11nes of investigation have suggested that Hultiple Sclerosis (MS) may be related to a chronic measles-like infection. We have investigated Natural Ktller (ilK) ce]] function from peripheral blood 1~mphocytes of 20 patients with MS and 20 age and sex matched volunteer controls against 3 cell lines: Hela; Measles infected Hela (11-Hela); K 56Z myeloid line; usin9 a chromium release mtcro-as-~ay. High levels of NK tn both I~S patients and controls were present against K 562 cell line. HLA B7 or DR2 was present tn 50~ of pati(nts vs. 25~ controls, p,.07o The haplotype A3,87 was found in 7/20 MS vs. 1/20 controls p<.02. I~K against N-Hela cell ltne was decreased tn MS patients compared to controls (MS 23+3~ vs° controls 34÷5Z specific chromt~ release). The decrease NK tn ftS"appeared most closely associated with A3,B7 haplotype (PIS-A3,B7 pus. vs. PIS*A3,B7 new., 145 vs. 28~: controls 32-475 p~.02). In contrast, there was no difference tn NK activity among the ~ patients with or without HLA-DR2 (10 DRZ pus., 10 DR2 neg.J, cur were there any differences ]n the number of circulating NK cells (Fc receptor positive for ]gG). In addition, though serum levels of measles antibody were higher tn MS than contrels, there was no correlation between NK activity against M-Hela and the level of measles antibody. In summary. not only ts there a selective decrease in NK activity of I~S patients against M-He]a, but this decrease ts most obvious tn patients possessing the A3,B7 haplotype. Thus, a deficiency tn the ]mmune surveillance capacity of SUSceptible individuals to a measles-like infection may contribute to the pathngenesis of Multiple Sclerosis. INHIBITION OF HELPERT CELL FUNCTIONNiD EXFRE$SIONOF "la" BY IIISTAHINE IND~ED SUPPRESSORFACTOR,H.R..GaroVOyo ~I.A. Reddisht R.E. l~ocklqn, Departments of Medicine Brigham and Women's Hospital and Tufts-New England I~dical Center, Boston, P;~. The effect of Histamine-Induced Suppressor Factor (HSF) on the humoral tmmune resoonse was examined wtth the mod~l of polyclonal B cell activat;on induced during a mixed l~,phecyte culture {MLC). TI~- number of p|aque forming ce]ls (PFC) g~nerated durinq IILC was measured by a Protein A plaque assay. HSF was produced by tncobatqn9 lymphocytes from norms] subjects ~tth 10- H histamine. Addition of HSF on day O, ~o IvLC induced plaques (measured on day 6) reduced the mean number of PFC/]O t e l | s : 1gG-1325, 1~-3725 and
Abstracts
36:~
I,A-1800 by G0-805 (4 expts). HSF suoernatants were active at a t t t e r of ]/1000 and suppressed lgG, IgH and lgA ~sotypes eque]]yo To study the effect of hSF on the T he|per cell component of th~s reaction, purified T |ymphocyteb were act~ ~ated ~n und~rect~ona] HLC (T*) and subsequently c ~ btned w~th unprimed B cel|s to induce a polyclona| PFC response. HSF present only during the qenerat~on phase of T*. or only at the time of coculture (T* ÷B). i,Mb~ted the total PFC response by 83 + ]3~ and 76 ~ 23~,o respectively (mean + $.D.. 3 expts). The expression of "la ~ and autoloqous DR antigens were no'-mallv detected on 50-62~ of T* generated duringMLC and reproducibly reduced to 20~ in the presence of HSF. Thus HSF inhibits MLC induced polycloaa| B ce]| activation by suppressing the 9eneration and function of helper T ce||s. The reduced expression of la/DR antigens may serve to ]~m~t the T-B cooperation required for a PFC response. MITOCEN-INDUCED PLASMA CELL DIFFERENTIATION IN PATIENTS ~TH MULTIPLE oLL~,u$1S EVIDENCE SUPPORTING A T CELL DEFECT. Howard M. Cebel. Leatha Ross. ~shn Trotter and Clenn E~Rodey~ Washillgton University School of Medicine, Departnents of Pathology and Neurology , S t . Louis, Missouri P e r i p h e r a l blood ly~phocytes from 9 p a t i e n t s w i t h a c t i v e l y p r o g r e s s i n g r ~ l t l p l e s c l e r o s i s (MS) o r normal v o l u n t e e r s were s e p a r a t e d i n t o T (>93Z p u r i t y ) and Non-T (40-58~ s u r f a c e I~ p o s l c x v e B c e l l s ) f r a c t i o n s by sheep ~rythrocyte rosetting C u l t u r e s co~calning I) T+B ( a u t o l o g o u s ; normal o r MS), 2) T+B (allogeneic; normal) 3) THs+BNor and ~) TNor+BMS were incubated fol seven d a y s z i n the p r e s e n c e of pokeweed mltogen (Pt~M). Each c u l t u r e c o n t a i n e d 5xlO ~ T c e l l s and 5xl0 ~ B c e l l s . The p e r c e n t a g e o f PWM induced plasma c e l l s wa~ determined by f l u o r e s c e n t s t a i n i n g f o r cytoplasmlc i,=,unoglobulin. When T+B c e i l s were c u l t u r e d a u t o l o g o u s l y from normal~ o r MS p a t i e n t s t h e r e was no d i f f e r e n c e i n the number of plasma c e U s g e n e r a t e d (21Z±4~ and 20.4Z±4.SZ, r e s p e c t i v e l y ) . In c o n t r a s t , a l l o g e n e i c c o m b i n a t i o n s ~f normal 5 + T c e l l s g e n e r a t e d 9 I*./I~.2Z plasma c e l l s . Allogeneic combinations o f TNor+BMS g e n e r a t e d 8.1Z±I.2Z plasma c e l l s w h i l e a l l o g e n e l c combinations o f TMS+BNer g e n e r a t e d 18.~Z±2.8Z plasma c e l l s (p< O.O01). I f TNo~ c e l l s were i r r a d i a t e d (900 RADS) b e f o r e c u l t u r l n g w i t h BMS c e l l s , t h e p e r c e n t a g e o f plasma c e l l s g e n e r a t e d was 21.I~±~.7Z. Our r e s u l t s s u g g e s t a c e l l a b n o r m a l i t y ( e . g . , a d e f e c t in s u p p r e s s o r o r c y t o t o x i c p r e c u r s o r s ) In p a t i e n t s w l t h M S . (Supported in p a r t by NIH g r a n t N$15977)
D~ONSTRATION OF IIUMAN T CFLL HETEROGENEITY. N.E..Goeken and J S. Thom~son~ Univ. of Iowa, Unlv. of Kentucky and VA Medical Centers. Theophylllne (Th), a phosphodzesterase inhibitor, also reverslb]y inhibits the SRBCreceptor of a subpopulatlon %~ human T lymphocytes.'Wehave prevlously reported that the precursor for the Concanaw/In A (Con ~) inducible buppressor ¢ell (SC) is found predominantly in the Th sensitive (S) populatlon, whereas the mltomyoxn resistant MLR induced SC is predominantly in the Threslstant (R) subset. Further studies now 3ndlcate addltiona~ l'unctlonal d i s t i n c t i o n s . The p r o l z f e r a t z v e an d c y t o t o x z e r e s p o n s e s o f T h - S a n i T h - R c e l l s t o a l l o g e n e t e s t i m u l a t i o n a r e approximately e q u a l , (h3,000 v s . 29,000 cpm; 32% v s . h0% s p e c i f i c 51Cr r e l e a s e ) a s are t h e r e s p o n s e s ~o PHA (39,000 v s . 29,000 olin). The r e s p o n s e t o Con A and pokeweed mxtogen i s up t o 20 Cold h i g h e r xn t h e Th-S p o p u l a t i o n s , however (12,000 v s . 900 cpm; 21,OOOvs° 1,200 cpm). Both c e l l t y p e s respond moderately (up t o 10,000 cpm) i n a u t o IogousMLR (AMLR), but d i f f e r zn their SC response to thls stimulus. ~ - R c e l l s a c t i v a t e d by I s o l a t e d , autologous non-T c e l l s in bulk c u l t u r e f o r 7 days a r e up t o h0%more s u p p r e s s i v e i n a second a l l o g e n e z e MLR a s s a y t h a n a r e Th-S c e l l s s i m i l a r l y t r e a t e d . AMLRInduced S C a r e completely mlto=~cin s e n s i t i v e , s u g g e s t i n g t h a t t h e MLR-SC and t h e AMLR-SC a r e f u n c t i o n a l l y d i f f e r e n t . Although t h e Th-S p o p u l a t i o n xs e n r i c h e d f o r Fcy+ c e l l s and Th-R f o r }¢~÷ c e l l s , c o r r e l a t i o n v~th t h e p r e v i o u s l y r e p o r t e d l"unct~onal p r o p e r t i e s o f c e l l s b e a r i n g t h o s e s u r f a c e phenotypes ~s not a b s o l u t e . These da~a amplify t h e concept t h a t T c e l l f u n c t i o n s , even " n o n - s p e c i f i c " o n e s , a r e not u b i q u i t o u s l y d i s t r i b u t e d , but r a t h e r , are c o n f i n e d t o d i s t i n c t but z n t e r d e pendant s u b s e t s . (Supported by t h e VA Research S e r v i c e ) .
Annual A A C H T Meeung, 1981
SPECIES CROSS- REACTIVE I a DETERMINANTS. DETECTION BY MONOCLONALANTIBODY TO HLT)La~ I s . JUDY FALK AND HICHELLE LETARTE, TORONTO LW.S,~E~ HOSPITAL AND HOSPITAL FOR S1CK CHILDREN, TORONTO, CANADA. A.TH a n t i - A . T L s e r m l , d i r e c t e d a t t h e I a a n t l g e n s o f murine h a p l o t y p e k . r e a c t s s t r o n g l y v i t h nen-polymorphic d e t e r m i n a n t s on a!1 huemn B c e l l s and B - C L L c e l l s t e s t e d . I a J and B p o l y p e p t l d e c h a i n s can be Immunoprecipltated from c e l l e x t r a c t s by t h e murine I-E/C s u b r e g i o n , ge have nov d e m o n s t r a t e d ~hat A.TH s n t l - A . T L s e r u m r e a c t s w l t h ~heep, p l g and r a t c e l l s a s ~ e l l a s v l t h human c e l l s . I n h i b i t i o n s ~ u g i e s , u s i n g t h e c e l l u l a r radlolmmunoassay, have d e m o n s t r a t e d t h a t s h e e p , p l g and r a t c e l l s c o u l d a b s o r b t h e b i n d i n g o f ~he A . T H a n t i - A . T L a e r u m t o htman I s . Thus t h i s " a l l o s e r u m '~ can a l s o r e c o g n i z e and I s d e t e r m i n a n t ( s ) common t o s e v e r a l s p e c i e s . A monoclonal a n t i b o d y , 21w4, r s l s e d b} I m u n l z l n g A.THmlce w i t h p u r i f i e d human l a g l y c o p r o t e i n s , d e t e c t e d nonpolymorphlc h e , a n l a d e t e t m l n a n t s I n t h e c e l l u l a r radiole~aunoassay; 21w4 a n t i b o d y a l s o r e a c t e d w i t h p i g , sheep and some mouse s t r a i n s . The murine s p e c i f i c i t y r e c o g n i z e d by t b i s monoclonal a n t i b o d y t o human I a m a p p o d t o t h e I-E/C s u b r e g i o n . The murine r e a c t i v i t y o f 21w4 c o u l d be a b s o r b e d by human, s h e e p and p i g c e l l s . T h e r e f o r e Immunization o f mtee of h a p l o t y p e s ( l a c k i n g I - E / C gene p r o d u c t s ) u i t h e l t h e r m u r l n e o r human l a m o l e c u l e s , e l i c i t s a n t i b o d i e s recognlzingamlno a c l d sequences conserve.l t h r o u g h o u t e v o l u t i o n , lqonoclonal a n t l b o d l ~ s such a s 21w4 a r e t h u s n o t o n l y v a l u a b l e f o r t h e puri~leatlonof human I s m o l e c u l e s b u t a l s o f o r t h e c h a r a c t e r l z a t l o n and p u r i f i c a t i o n o f I s m o l e c u l e s from s e v e r a l s p e c i e s .
THE A3,B7 HAPLOTYPE. DECREASEDNATURALKILLER (NK) CELL ACTIVITY IN MULTIPLE SCLEROSIS, H.R._Garovoyo $. Hau~er~ K. AulL. C. Connors~ H.L. We]net. Depts. of Medicine and Neurology, Brigham and Women's Hospttalo Boston, Has-s. Several 11nes of investigation have suggested that Hultiple Sclerosis (MS) may be related to a chronic measles-like infection. We have investigated Natural Ktller (ilK) ce]] function from peripheral blood 1~mphocytes of 20 patients with MS and 20 age and sex matched volunteer controls against 3 cell lines: Hela; Measles infected Hela (11-Hela); K 56Z myeloid line; usin9 a chromium release mtcro-as-~ay. High levels of NK tn both I~S patients and controls were present against K 562 cell line. HLA B7 or DR2 was present tn 50~ of pati(nts vs. 25~ controls, p,.07o The haplotype A3,87 was found in 7/20 MS vs. 1/20 controls p<.02. I~K against N-Hela cell ltne was decreased tn MS patients compared to controls (MS 23+3~ vs° controls 34÷5Z specific chromt~ release). The decrease NK tn ftS"appeared most closely associated with A3,B7 haplotype (PIS-A3,B7 pus. vs. PIS*A3,B7 new., 145 vs. 28~: controls 32-475 p~.02). In contrast, there was no difference tn NK activity among the ~ patients with or without HLA-DR2 (10 DRZ pus., 10 DR2 neg.J, cur were there any differences ]n the number of circulating NK cells (Fc receptor positive for ]gG). In addition, though serum levels of measles antibody were higher tn MS than contrels, there was no correlation between NK activity against M-Hela and the level of measles antibody. In summary. not only ts there a selective decrease in NK activity of I~S patients against M-He]a, but this decrease ts most obvious tn patients possessing the A3,B7 haplotype. Thus, a deficiency tn the ]mmune surveillance capacity of SUSceptible individuals to a measles-like infection may contribute to the pathngenesis of Multiple Sclerosis. INHIBITION OF HELPERT CELL FUNCTIONNiD EXFRE$SIONOF "la" BY IIISTAHINE IND~ED SUPPRESSORFACTOR,H.R..GaroVOyo ~I.A. Reddisht R.E. l~ocklqn, Departments of Medicine Brigham and Women's Hospital and Tufts-New England I~dical Center, Boston, P;~. The effect of Histamine-Induced Suppressor Factor (HSF) on the humoral tmmune resoonse was examined wtth the mod~l of polyclonal B cell activat;on induced during a mixed l~,phecyte culture {MLC). TI~- number of p|aque forming ce]ls (PFC) g~nerated durinq IILC was measured by a Protein A plaque assay. HSF was produced by tncobatqn9 lymphocytes from norms] subjects ~tth 10- H histamine. Addition of HSF on day O, ~o IvLC induced plaques (measured on day 6) reduced the mean number of PFC/]O t e l | s : 1gG-1325, 1~-3725 and
Abstracts
36:~
I,A-1800 by G0-805 (4 expts). HSF suoernatants were active at a t t t e r of ]/1000 and suppressed lgG, IgH and lgA ~sotypes eque]]yo To study the effect of hSF on the T he|per cell component of th~s reaction, purified T |ymphocyteb were act~ ~ated ~n und~rect~ona] HLC (T*) and subsequently c ~ btned w~th unprimed B cel|s to induce a polyclona| PFC response. HSF present only during the qenerat~on phase of T*. or only at the time of coculture (T* ÷B). i,Mb~ted the total PFC response by 83 + ]3~ and 76 ~ 23~,o respectively (mean + $.D.. 3 expts). The expression of "la ~ and autoloqous DR antigens were no'-mallv detected on 50-62~ of T* generated duringMLC and reproducibly reduced to 20~ in the presence of HSF. Thus HSF inhibits MLC induced polycloaa| B ce]| activation by suppressing the 9eneration and function of helper T ce||s. The reduced expression of la/DR antigens may serve to ]~m~t the T-B cooperation required for a PFC response. MITOCEN-INDUCED PLASMA CELL DIFFERENTIATION IN PATIENTS ~TH MULTIPLE oLL~,u$1S EVIDENCE SUPPORTING A T CELL DEFECT. Howard M. Cebel. Leatha Ross. ~shn Trotter and Clenn E~Rodey~ Washillgton University School of Medicine, Departnents of Pathology and Neurology , S t . Louis, Missouri P e r i p h e r a l blood ly~phocytes from 9 p a t i e n t s w i t h a c t i v e l y p r o g r e s s i n g r ~ l t l p l e s c l e r o s i s (MS) o r normal v o l u n t e e r s were s e p a r a t e d i n t o T (>93Z p u r i t y ) and Non-T (40-58~ s u r f a c e I~ p o s l c x v e B c e l l s ) f r a c t i o n s by sheep ~rythrocyte rosetting C u l t u r e s co~calning I) T+B ( a u t o l o g o u s ; normal o r MS), 2) T+B (allogeneic; normal) 3) THs+BNor and ~) TNor+BMS were incubated fol seven d a y s z i n the p r e s e n c e of pokeweed mltogen (Pt~M). Each c u l t u r e c o n t a i n e d 5xlO ~ T c e l l s and 5xl0 ~ B c e l l s . The p e r c e n t a g e o f PWM induced plasma c e l l s wa~ determined by f l u o r e s c e n t s t a i n i n g f o r cytoplasmlc i,=,unoglobulin. When T+B c e i l s were c u l t u r e d a u t o l o g o u s l y from normal~ o r MS p a t i e n t s t h e r e was no d i f f e r e n c e i n the number of plasma c e U s g e n e r a t e d (21Z±4~ and 20.4Z±4.SZ, r e s p e c t i v e l y ) . In c o n t r a s t , a l l o g e n e i c c o m b i n a t i o n s ~f normal 5 + T c e l l s g e n e r a t e d 9 I*./I~.2Z plasma c e l l s . Allogeneic combinations o f TNor+BMS g e n e r a t e d 8.1Z±I.2Z plasma c e l l s w h i l e a l l o g e n e l c combinations o f TMS+BNer g e n e r a t e d 18.~Z±2.8Z plasma c e l l s (p< O.O01). I f TNo~ c e l l s were i r r a d i a t e d (900 RADS) b e f o r e c u l t u r l n g w i t h BMS c e l l s , t h e p e r c e n t a g e o f plasma c e l l s g e n e r a t e d was 21.I~±~.7Z. Our r e s u l t s s u g g e s t a c e l l a b n o r m a l i t y ( e . g . , a d e f e c t in s u p p r e s s o r o r c y t o t o x i c p r e c u r s o r s ) In p a t i e n t s w l t h M S . (Supported in p a r t by NIH g r a n t N$15977)
D~ONSTRATION OF IIUMAN T CFLL HETEROGENEITY. N.E..Goeken and J S. Thom~son~ Univ. of Iowa, Unlv. of Kentucky and VA Medical Centers. Theophylllne (Th), a phosphodzesterase inhibitor, also reverslb]y inhibits the SRBCreceptor of a subpopulatlon %~ human T lymphocytes.'Wehave prevlously reported that the precursor for the Concanaw/In A (Con ~) inducible buppressor ¢ell (SC) is found predominantly in the Th sensitive (S) populatlon, whereas the mltomyoxn resistant MLR induced SC is predominantly in the Threslstant (R) subset. Further studies now 3ndlcate addltiona~ l'unctlonal d i s t i n c t i o n s . The p r o l z f e r a t z v e an d c y t o t o x z e r e s p o n s e s o f T h - S a n i T h - R c e l l s t o a l l o g e n e t e s t i m u l a t i o n a r e approximately e q u a l , (h3,000 v s . 29,000 cpm; 32% v s . h0% s p e c i f i c 51Cr r e l e a s e ) a s are t h e r e s p o n s e s ~o PHA (39,000 v s . 29,000 olin). The r e s p o n s e t o Con A and pokeweed mxtogen i s up t o 20 Cold h i g h e r xn t h e Th-S p o p u l a t i o n s , however (12,000 v s . 900 cpm; 21,OOOvs° 1,200 cpm). Both c e l l t y p e s respond moderately (up t o 10,000 cpm) i n a u t o IogousMLR (AMLR), but d i f f e r zn their SC response to thls stimulus. ~ - R c e l l s a c t i v a t e d by I s o l a t e d , autologous non-T c e l l s in bulk c u l t u r e f o r 7 days a r e up t o h0%more s u p p r e s s i v e i n a second a l l o g e n e z e MLR a s s a y t h a n a r e Th-S c e l l s s i m i l a r l y t r e a t e d . AMLRInduced S C a r e completely mlto=~cin s e n s i t i v e , s u g g e s t i n g t h a t t h e MLR-SC and t h e AMLR-SC a r e f u n c t i o n a l l y d i f f e r e n t . Although t h e Th-S p o p u l a t i o n xs e n r i c h e d f o r Fcy+ c e l l s and Th-R f o r }¢~÷ c e l l s , c o r r e l a t i o n v~th t h e p r e v i o u s l y r e p o r t e d l"unct~onal p r o p e r t i e s o f c e l l s b e a r i n g t h o s e s u r f a c e phenotypes ~s not a b s o l u t e . These da~a amplify t h e concept t h a t T c e l l f u n c t i o n s , even " n o n - s p e c i f i c " o n e s , a r e not u b i q u i t o u s l y d i s t r i b u t e d , but r a t h e r , are c o n f i n e d t o d i s t i n c t but z n t e r d e pendant s u b s e t s . (Supported by t h e VA Research S e r v i c e ) .