of Anaphylactic Shock: Studies on Complement

Transcription

1 Journl of inicl Investigtion Vol. 43, No. 4, 194 The Initil Stge of Cnine Endotoxin Shock s n Expression of Anphylctic Shock: Studies on Complement Titers nd Plsm Histmine Concentrtions * WESLEY W. SPINK, RICHARD B. DAVIS,t RUTH POTTER, AND SANDRA CHARTRAND (From the Deprtment of Medicine, University of Minnesot Hospitls nd Medicl School, Minnepolis, Minn.) Within 3 to seconds fter endotoxin is injected into dult mongrel dogs there is decline in systemic blood pressure, rise in portl vein pressure, reduction in the venous return of blood to the hert, nd decrese in renl blood flow (1, 2). Endotoxin shock is comprble in mny wys to cute nphylctic shock in the dog (3). Severl observtions hve suggested tht n immune mechnism cuses the initil vsculr chnges. It ws demonstrted tht the ction of endotoxin on blood vessels ws medited through het lbile fctor in plsm or serum (4, 5). Gilbert nd Brude () in studies on Escherichi coli endotoxin shock in rbbits reported tht doses of endotoxin greter thn the LD5 cused decline in the titer of complement nd decrese in the serum concentrtion of E. coli ntibody. Spink nd Potter (7) lso observed prompt decrese in plsm complement vlues in cnine endotoxin shock. The immedite effect of endotoxin on smller vessels suggested tht vsoctive substnce or substnces were lso implicted. If n immune mechnism were involved, histmine could be one of the substnces tht ws liberted by n ntigenntibody mechnism, nd studies do point to histmine s fctor cusing the ltered vsculr ctivity (3, 8-1). This report concerns the study of complement titers, plsm histmine concentrtions, nd blood pressure chnges in series of dogs given lethl * Submitted for publiction August 15, 193; ccepted December 12, 193. Supported by U. S. Public Helth Service grnt AI t Recipient of U. S. Public Helth Service Reserch Creer Progrm Awrd 5-K3-HE from the Ntionl Hert Institute. dose of endotoxin. In ddition, since epsilonminocproic cid nd cortisol protect dogs ginst endotoxin, we were interested in scertining the effect of these two gents upon complement nd histmine vlues. The dt support the concept tht the initil hemodynmic phse of cnine endotoxin shock is relted to n immune mechnism of the immedite nphylctic type. 9 Methods Animls. A totl of 85 dult mongrel dogs ws used in series of 1 experiments. Endotoxin. The sme lot of E. coli endotoxin ws used throughout nd ws prepred s described elsewhere (11). The LD1 of the endotoxin ws first estblished in mice nd then in dogs. The LD,., for the ltter species ws.55 mg per kg. Complement ssy. Titers were expressed s 5% hemolytic units, employing the method outlined by Kbt nd Myer (12). Blood ws collected before the injection of endotoxin nd then 1 minutes, 1, 3, nd hours lter. After the blood hd clotted t room temperture, serum ws removed by centrifugtion nd stored t 2 - C overnight. Control studies showed tht this temporry storge did not result in significnt deteriortion of complement. Histmine ssy. Seril plsm histmine concentrtions were determined by modifiction (13) of the fluorometric ssy described by Shore, Burkhlter, nd Cohn (14). The relibility of this method ws determined by biossy on duplicte smples through the courtesy of Dr. Chrles Code of the Myo Foundtion for Medicl Reserch. A good correltion of the two techniques ws found. Epsilon-minocproic cid (EACA).1 One g contined in sterile vils ws infused intrvenously in 1 ml of 5% dextrose nd distilled wter over period of 15 minutes before the dministrtion of endotoxin. Complement nd histmine ssys were crried out before nd fter the injection of EACA, nd fter endotoxin hd been given. 1 Supplied by Lederle Lbortories, Americn Cynmid Co., Perl River, N. Y.

2 ANAPHYLAXIS AND CANINE ENDOTOXIN SHOCK 97 Cortisol.2 One hundred mg contined in sterile vils ws infused intrvenously in 1 ml of 5%o dextrose nd wter over period of 15 minutes before the dministrtion of endotoxin. Complement nd histmine ssys were likewise obtined fter this infusion, s well s fter endotoxin. EACA plus cortisol. Quntities of these two gents s given bove were infused simultneously into nimls, nd complement nd histmine vlues were obtined during the sme periods before nd fter the injection of endotoxin. Observtions on shock. The nimls, nesthetized with sodium pentobrbitl, were studied in mnner previously described (1). Continuous femorl rteril blood pressure determintions nd urinry output were recorded up until hours fter endotoxin, or until deth. Control nimls. To ssess the role of nesthesi, the loss of blood necessry for the tests, trum due to femorl ctheteriztion, nd the effect of intrvenous injections of physiologic solutions, the following studies were crried out: 1) complement titers on three dogs nesthetized nd given 1 ml of 5% dextrose nd wter (this ws done to control the effects on complement following the infusion of EACA nd cortisol); 2) seril histmine ssys on six dogs to determine the effects of nesthesi, the trum of ctheteriztion, nd the injection of 2 ml of sline, which ws the mount of solution required for the dministrtion of endotoxin; 3) complement titers in six dogs fter lethl dose of the snke venom, Crotlus terrificus,3 since the initil hemodynmic ltertions in the dog could be produced by substnces other thn-endotoxin, such s snke venom. Results 1. Complement titers nd histmine concentrtions in dogs given lethl dose of endotoxin. 2 Supplied s Solu-Cortef (hydrocortisone sodium succinte) by Upjohn Co., Klmzoo, Mich. 3 Supplied by University of So Pulo, So Pulo, Brzil. z W 1 w E 8 (-) o (I4 z 4 (158) PRE ENDO. Min. -PPOST ENDOTOXIN FIG. 1. DECREASE IN COMPLEMENT TITERS IN EACH OF TEN DOGS GIVEN.55 MG PER KG ENDOTOXIN. There were no survivors. Ten dogs, weighing from 8.1 to 12.5 kg, were nesthetized nd given.55 mg per kg of endotoxin. Survivl times vried from 2- to 1 hours. The decline in complement is shown grphiclly in Figure 1. Preliminry studies on complement titers t 1, 5, 1, nd 3 minutes fter lethl dose of endotoxin hd shown tht the verge mximl decline occurred t 1 minutes. After this period there ws tendency for the complement titers to rise, especilly in dogs tht recovered. Although rbitrry, the time of pprent mximl decline of complement ws chosen for comprison in most studies. The decline of complement in nimls 1 minutes fter endotoxin (Tble I) TABLE I Percentge of chnge in complement titer in dogs 1 minutes fter lethl dose of endotoxin (.55 mg/kg)* Pretreted Survivors Control, Pretreted Pretreted with given 2nd 5% Control, with with EACA dose dextrose endotoxin EACA cortisol nd cortisol endotoxin Men i SE + i * EACA = epsilon-minocproic cid.

3 98 * - ' *' I 4 *. U '. ~w to- lc- IC 4) U to c- 8 E - v o A u UW@ 4) 8. W = I...i v U ~In 4 :r (n LI) VI) v r.y 5`_ I cn :: :4 (n W. W. SPINK, R. B. DAVIS, R. POTTER, AND S. CHARTRAND t-. en - o n C\ o 4 o en CN CN C1 o o) o o I- t- 'I It C ) C I " - - Uf) C\ (DI C \ en so m ~o cl m C> c"i eq cq I -4 4 en -4 ' 5 - N f) C on en o - - \o en o o en " q en U) t- o o C% c )-4 1- N- n mt o oo N - r-u) CUl Ci C) o o. o o- C' ) UO) t- o o " en) ' Itt " N *; Uf) '- -4 t) - t- - Ci O )o C I- If) (f W 4 cd Q U) ) 4)x. * * ws highly significnt compred to sline control nimls (p <.1). Plsm histmine concentrtions re presented in Tble II. Mny preliminry observtions reveled tht the mximl rise in plsm histmine occurred between 3 nd seconds fter the injection of endotoxin. An explosive rise occurred in five dogs t the end of seconds, reveling n verge concentrtion 8 times the vlue obtined before endotoxin ws injected. This elevtion ws lso confirmed in selected nimls by the biossy method used by Dr. Code. The men increse in plsm histmine ws significntly greter thn in sline-injected dogs 5 minutes fter endotoxin (p <.5), but ws not sttisticlly significnt fter 3 nd seconds. In dogs tht survived 4 hours, histmine vlues pproched those of the control pre-endotoxin period. It hs been repetedly observed in the dog tht n brupt drop in systemic pressure occurred 1 to 5 minutes fter endotoxin. Within 15 to 3 minutes pressures usully pproched the pre-endotoxin bse line. There ws good correltion between the brupt rise in histmine nd the initil period of hypotension (Tble II), lthough two nimls filed to show n pprecible increse in histmine concentrtions, nd in three initil hypotension preceded demonstrted increse in histmine concentrtions. 2. Complement titers in dogs given n LD5,, injection of endotoxin. Six nimls given n o LD5o injection reveled distinct difference in the vlues of complement on the bsis of survivl or deth s seen in Figure 2. All nimls hd m4 n initil drop in complement. In the three nir mls tht survived the men titer ws 77%o of 1 the control, wheres in the three tht expired the, men titer ws 23%o of control fter 3 hours. Cd This difference is highly significnt (p <.1). 3. Complement titers nd histmine concen- Cd trtions in dogs pretreted -with EACA nd then given lethl dose of endotoxin. In previous o. studies the mjority of dogs pretreted with n EACA survived lethl dose of endotoxin (15). Ten dogs were given 1 g of EACA s n infusion '< in 1 ml of 5% dextrose nd wter. At the end * of this time.55 mg per kg of endotoxin ws injected. Seven of the 1 dogs survived.

4 ANAPHYLAXIS AND CANINE ENDOTOXIN SHOCK Complement titers in this group of nimls were not pprecibly ltered by the initil infusion of 1 g of EACA in 1 ml of 5% dextrose nd wter, the men decrese in complement being 19%o in nimls receiving dextrose nd wter lone, nd 25%o in those given EACA in dextrose nd wter. The men complement titer tht ws present fter the EACA infusion nd before the injection of endotoxin ws not significntly different from the vluies in those control nimls receiving endotoxin lone. In the dogs pretreted with EACA there ws no difference in the fll of complement fter 1 minutes in those tht survived nd those tht died. Men plsm histmine concentrtions of 1 nimls pretreted with EACA nd then given lethl dose of endotoxin re shown in Tble II. The verge vlues for this treted group re only slightly lower thn the untreted control group. Coincident with the rise in plsm histmine there ws moderte fll in blood pressure. The infusion of EACA lone cuses rise in systemic pressure, n observtion tht hs been confirmed by others (1). This experiment showed tht pretretment of dogs with EACA significntly reduced the mortlity rte from endotoxin nd lso reduced the fll in complement when compred to nimls receiving endotoxin lone (p <.2), s noted in Tble I. In EACA-treted nimls plsm histmine concentrtions rose significntly seconds fter endotoxin (p <.5). 4. Complement titers nd histmine concentrtions in dogs pretreted with cortisol nd then given lethl dose of endotoxin. Pretretment with cortisol protects dogs ginst lethl dose of endotoxin. Ten dogs were given 1 mg of cortisol intrvenously in 1 ml of 57o dextrose nd sline. Seven of the 1 dogs survived. As seen in Tble I this mount of cortisol did not significntly lter the initil decline in complement titers 1 minutes fter endotoxin, s compred to the titers in control endotoxin nimls (p =.2). The men pre-endotoxin titer fter cortisol infusion ( U) ws lower thn in endotoxin-treted dogs (p <.1). Cortisol hs pronounced effect in suppressing the rise in plsm histmine concentrtions resulting from endotoxin (Tble II). The control z 1 w w (] 8, u Ded --- Survivor 99 PRE F ENDO MPTin -POST ENDOTOXIN FIG. 2. DECREASE IN COMPLEMENT TITERS IN EACH OF SIX DOGS GIVEN LD3 DOSE OF ENDOTOXIN (.25 MG PER KG). The decline in complement ws less in the survivors thn in those tht died. nimls receiving endotoxin lone hd n eightfold rise, wheres nimls pretreted with cortisol hd slightly less thn fourfold elevtion. In nimls pretreted with cortisol nd given endotoxin the plsm histmine concentrtions were no greter thn in those nimls given sline solution lone. In ddition, the initil fll in systemic pressure ws somewht less in cortisoltreted nimls thn in the control group. Sixty seconds fter injection of endotoxin, when the mximl verge level of.19 ug per ml of histmine ws reched in the plsm, the verge systolic blood pressure ws 13 mm Hg. Comprble vlues in the control endotoxin group were.4 Mig per ml of histmine nd pressure of 91 mm Hg. Although the differences in plsm histmine were not sttisticlly significnt fter 3 nd seconds, 5 minutes fter endotoxin plsm histmine in the cortisol-treted group ws.9 Mtg per ml, nd in the endotoxin controls,.27; this difference ws significnt (p <.2). The respective men systolic blood pressures were 123 nd 95 (Tble II). 5. Complement titers nd histmine concentrtions in dogs pretreted zwith both EACA nd cortisol nd then given lethl dose of endotoxin. Studies reported elsewhere (15, 17) indicted tht different mechnisms were involved for EACA nd for cortisol in the protection fforded dogs ginst endotoxin. For this reson 1 dogs were

5 7 W. W. SPINK, R. B. DAVIS, R. POTTER, AND S. CHARTRAND, % - o c - % o 11) (n otm o 1)m O O O l 1 V) ~~~~~~~~~C *r; *; I' 41.4 cx; (14 CU" ONc; t U'- 1f) c1) % C')) Oto oo U'. ) E f< c v. n U) CS *o ; 4.. 1') c '1) 11 U'- Cō > CU- Xf)~~~~~~~~~Y % U) *) r Cx; C; U'. 11) Co4 o o i oo OO ri. > CdC IC b4 41 bo x. )z i o. in) -4 o r o 4 1- o o v oo Xf n4 C' C') 1c) _ U o o 1i uoc) 1.) *.I - * U. U. c; CN 1p o 1, \ )~~~~~~~~~C.U> C.) Cd U I- c)." \ - M-4 C' I 11 X CU Cx; Co 11) Ol o- I- H 11o) oo o ) ooo ) U). o r' o o.g4 I,) C >- - U) ) o o U U) > Ul) ~~~~' cd CE - cd c cd Cd E2E E8 3-. (n I r m CO ~~~~~ CdU 'u 4 11)~ 4 cu W~*J). 4 ) U)). CU C) E cu ) I.1,11 U) U2 cu. 1: ~,.M 'I *

6 ANAPHYLAXIS AND CANINE ENDOTOXIN SHOCK infused simultneously with 1 g EACA nd 1 mg cortisol contined in 1 ml 5% dextrose nd wter. Seven of 1 nimls survived.55 mg per kg of endotoxin, which ws the sme outcome in n equl number of nimls pretreted with EACA or cortisol lone. This combintion lone hd no effect on the complement titer compred to dextrose-infused dogs. The men control titer fter EACA nd cortisol (87 U) did not differ significntly from the endotoxin control group (93 U). Ten minutes fter the injection of endotoxin the verge decrese in titer ws less thn tht observed in control endotoxin nimls (p <.1) or in those pretreted with either EACA or cortisol (Tble I) Ṫhe verge concentrtion of plsm histmine in this group did not exceed.9 pg per ml (Tble II). Five minutes fter endotoxin the men plsm histmine concentrtion ws less (.8) thn in nimls given only endotoxin (.27); the difference ws significnt (p <.2). It ws gin observed tht the systemic pressure rose fter the infusion of EACA nd cortisol, nd the verge level in systolic pressure fter endotoxin in this group did not decline below 1 mm Hg.. Complement titers nd histmine concentrtions in dogs given second lethl dose of endotoxin. A group of dogs treted previously by vriety of methods hd survived lethl dose of endotoxin (Tble III). After period of survivl vrying between few dys to over 3 months, second lethl dose of.55 mg per kg of endotoxin ws given to ech of these nimls. Although there ws n immedite nd severe rection following this second injection, eight of the ten nimls survived. There ws mrked decline in the titer of complement (Tble I), which exceeded tht in other groups nd ws significntly greter thn in nimls which received one dose of endotoxin (p <.5), but the decrese hd no reltion to the subsequent deth or survivl of the niml. The highest sustined concentrtions of histmine were observed in this group of nimls (Tble III). Within seconds fter endotoxin ws injected there ws 13-fold increse. In- 71 cresed concentrtions were present 4 hours lter in five nimls. The remrkble feture bout these nimls ws the prompt fll in complement, the onset of hypotension, nd mrked rise in plsm histmine, ll of which suggested n ntigen-ntibody rection of- the immedite nphylctic type. Nevertheless, eight of ten nimls recovered. 7. Histmine concentrtions in dogs pretreted with histmine nd then given lethl dose of endotoxin. Although dogs receiving the second dose of endotoxin exhibited the initil mnifesttions of severe nphylctic shock nd hd sustined increses of histmine, only two of the ten nimls died (Tble II). This suggested tht histmine might ctully protect nimls ginst the lethl effect of endotoxin, lthough one of its properties is the production of hypotension. Ten dult dogs were ech given n infusion of histmine phosphte, equivlent to 1 mg histmine bse, in 1 ml 5% dextrose nd wter over period of 15 to 2 minutes. Although some hypotension ws induced with histmine, the verge systolic blood pressure did not decrese below 9 mm Hg. After this infusion the nimls were then given lethl dose of endotoxin (.55 mg per kg). The initil rection to the endotoxin ws mild, nd eight of the ten nimls survived. There ws no rise in plsm histmine fter endotoxin (Tble II). 8. Control nimls. ) Complement titers in dogs given Cr. terrificus venom. Six dult nimls were given 5 mg of snke venom contined in.5 ml of sline. Although lethl shock ws produced in ll of the dogs the men complement titer ws 9%o of the control titer 1 minutes fter venom, nd chnged little therefter. b) Complement titers in nimls given 1 ml 5%o dextrose in distilled wter. Since EACA nd cortisol were given intrvenously in - solution of 5%o dextrose nd 1 ml of distilled wter, this mount ws infused into three nesthetized dogs. Complement titers fell pproximtely 19%o during the infusion of dextrose, but therefter complement chnged little, nd no chnge in blood pressure occurred. c) Plsm histmine concentrtions in dogs given 2 ml of sline solution intrvenously. After nesthesi, rteril ctheteriztion, nd iv injection of 2 ml

7 72 W. W. SPINK, R. B. DAVIS, R. POTTER, AND S. CHARTRAND of sline, plsm histmine ws mesured serilly in six dogs. No significnt chnges in concentrtion were obtined, with the men ( determintions) being.5 fug per ml. Blood pressure lwys fell in nimls given endotoxin, nd the fll ws ssocited with vrible rises in histmine nd decreses in complement titers. Nevertheless, clcultion of correltion coefficients between the mximl fll of blood pressure nd mximl chnge of histmine or complement in ech of the six groups showed no sttisticlly significnt correltion. Discussion The foregoing experiments were primrily concerned with the initil hemodynmic ltertions in the dog tht re induced by endotoxin. The dt support the concept tht the initil phse of cnine endotoxin shock involves n immune mechnism of the nphylctic type in which endotoxin, cting s n ntigen, rects with ntibody in the presence of complement, nd histmine is liberted. This concept does not exclude the ppernce of other vsoctive substnces in the blood. Other studies lso support such concept. Gilbert nd Brude () in immunologic studies on endotoxin shock in the rbbit concluded tht endotoxin hd n nphylctic effect. Kostk nd Sterzl (18) observed tht the serum of piglets removed from their mothers t birth contined complement but no E. coli ntibody. When endotoxin ws dded to piglet serum no inctivtion of complement occurred. However, the ddition of endotoxin to the ser of dult swine contining ntibody did result in significnt decrese in complement. In contrst to the evidence suggesting tht endotoxin cts s n ntigen in the initil phse of shock, Frnke (19) found tht lthough polyscchride from Serrti mrcescens ws lethl for guine pigs, the effects did not pper to be due to nphylxis. They mesured only lung volume nd specific grvity, however, nd it hs been shown in nother species tht mny objective signs of toxicity during nphylxis my be bsent despite the occurrence of chemicl chnges (2). There is considerble body of evidence indicting tht the initil chnges in vsculr ctivity in endotoxin shock cn be provoked by histmine. Unique in the dog is the immedite heptovenoconstriction with n increse in portl vein pressure tht occurs fter endotoxin nd is ssocited with pooling of blood long the intestinl trct. Essex nd Thoms (21) observed in dogs tht the intrvenous injection of histmine resulted in constriction of the heptic vein, nd Weil nd Spink (3) demonstrted prompt rise in histmine-like substnce in the heptic vein blood of dogs immeditely fter the injection of endotoxin. They were ble to demonstrte histmine-like ctivity in heptic vein blood but not in femorl vein blood, which is in ccord with the vrible increses in histmine found in this study in femorl rtery smples. Histmine cuses venoconstriction in the tissues nd orgns of other species (22, 23). Endotoxin produces venous constriction of pulmonry veins in the ct nd dog (24). Finlly, the hypoxi nd reduction of oxygen content in the dog's blood tht follows endotoxin hs been produced by histmine (25). While these studies re consistent with the hypothesis tht the erly phse of cnine endotoxin shock is relted to n ntigen-ntibody rection involving complement, resulting in histmine libertion, the precise role of histmine in the over-ll genesis of irreversible shock is not cler. Schyer (2) hs dvnced the thesis tht excessive mounts of histmine or epinephrine re deleterious to the microcircultion of the host. He hs concluded tht histmine ppers in the blood fter the onset of shock s result of the libertion of preformed histmine nd lso becuse of continued synthesis of histmine from histidine through the ction of the enzyme, histidine decrboxylse (27). The continued synthesis of histmine is more importnt in the genesis of shock thn is the libertion of preformed histmine. Others (1) support this dul origin of histmine, lthough Wton (28) questions the continuing synthesis of histmine in cts, dogs, nd mn. Since the present experiments re concerned with cute nd lethl endotoxin shock, no conclusive dt on the continued elevtion of plsm histmine over period of severl hours hve been obtined except in those surviving nimls given second injection of endotoxin. Severl provoctive points relting to the significnce of histmine in endotoxin shock hve been brought out in the present studies. First,

8 ANAPHYLAXIS AND CANINE ENDOTOXIN SHOCK 73 the highest sustined concentrtions of plsm histmine were detected in those dogs tht hd been given second lethl dose of endotoxin. This elevtion of histmine ws ccompnied by the mnifesttions of severe nphylctic shock. However, the mjority of these nimls survived. A stte of tolernce to endotoxin ws ssocited with cquired endotoxin hypersensitivity. The nture of this reltionship is not cler. It is difficult to correlte the initil severe nphylctic rection with the mechnism or mechnisms by which recovery ensues, if indeed, correltion exists. It is possible tht the stte of tolernce to the lethl effect of endotoxin is relted to the reticuloendothelil system, s the studies of others would suggest (29, 3). Second, dogs given n infusion of histmine over period of 15 minutes re rendered highly tolernt to the lethl effect of endotoxin, n observtion tht others hve mde in mice (31). An explntion for this phenomenon is not esily forthcoming. The injection of endotoxin ws not followed by significnt elevtions of plsm histmine fter histmine infusion. It is known tht histmine stimultes the secretion of epinephrine from the drenl medulle. Is it possible tht the injection of pproprite mounts of histmine depletes or diminishes the potentil secretion of epinephrine fter the injection of endotoxin? This spect is under investigtion. Third, EACA or cortisol protects dogs ginst lethl dose of endotoxin, lthough the mechnism by which ech of these gents ccomplishes this effect is probbly different. EACA but not cortisol pretretment significntly decresed the consumption of complement fter endotoxin, when compred to the results in control nimls (Tble I). The rise in plsm histmine from endotoxin ws no different in EACA pretreted nimls, but ws much less in cortisol-treted nimls thn in control nimls given endotoxin (Tble II). Libertion of histmine from cells is probbly inhibited by cortisol t the cell surfce, since the libertion of severl cellulr enzymes by endotoxin is considerbly diminished both in vitro nd in vivo in the presence of phrmcologic mounts of cortisol (32, 33). It is pprent tht endotoxin shock involves chin of biochemicl nd functionl ltertions. Becuse of this complexity shock cn be reversed t different stges nd by vriety of gents (34). Further progress in the understnding of this type of shock is dependent upon more precise chemicl identifiction of endotoxin. Additionl informtion is desirble on the quntittive reltionship of histmine nd ctecholmines during the vrious phses of shock. Other vsoctive substnces my lso be involved. Dt re especilly needed in different species, including mn. Summry Experimentl dt in cnine endotoxin shock support the concept tht the initil stge of hemodynmic ltertions is due to n nphylctic type of immune mechnism involving complement, with the libertion of histmine. However, the severity of the systemic rection is not indictive of the finl outcome of nimls. Epsilon-minocproic cid nd cortisol modify the severity of the initil rection, nd the mjority of nimls survive lethl doses of endotoxin. On the other hnd, surviving nimls given second lethl dose of endotoxin hve severe initil rection, significnt decline in complement, nd mrked rise in plsm histmine. The mjority of these nimls lso survive. Finlly, dogs infused with histmine survive lethl dose of endotoxin. The reltionship of the initil nphylctic ctivity to the ultimte outcome of n niml with endotoxin shock is not cler. Tolernce or intolernce to the lethl ction of endotoxin is dependent upon other mechnisms nd my be relted to cquired humorl immunity nd conditioning of the reticuloendothelil system. References 1. Weil, M. H., L. D. McLen, M. B. Visscher, nd W. W. Spink. Studies on the circultory chnges in the dog produced by endotoxin from grm-negtive microorgnisms. J. clin. Invest. 195, 35, Hinshw, L. B., W. W. Spink, J. A. Vick, E. Mllet, nd J. Finstd. Effect of endotoxin on kidney function nd renl hemodynmics in the dog. Amer. J. Physiol. 191, 21, Weil, M. H., nd W. W. Spink. A comprison of shock due to endotoxin with nphylctic shock. J. Lb. clin. Med. 1957, 5, Vick, J. Studies on the trigger mechnism involved in the vsculr ltertions induced by endotoxin. (bstrct). J. Lb. clin. Med. 19, 5, 953.

9 74 W. W. SPINK, R. B. DAVIS, R. POTTER, AND S. CHARTRAND 5. Spink, W. W., nd J. Vick. A lbile serum fctor in experimentl endotoxin shock: cross-trnsfusion studies in dogs. J. exp. Med. 191, 114, 51.. Gilbert, V. E., nd A. I. Brude. Reduction of serum complement in rbbits fter injection of endotoxin. J. exp. Med. 192, 11, Spink, W. W., nd R. Potter. Endotoxin shock nd immunologic fctors: studies on complement. Fed. Proc. 193, 22, Greismn, S. E. Activtion of histmine-relesing fctor in norml rt plsm by E. coli endotoxin. Proc. Soc. exp. Biol. (N. Y.) 19, 13, Hinshw, L. B., J. A. Vick, C. H. Crlson, nd Y-L. Fn. Role of histmine in endotoxin shock. Proc. Soc. exp. Biol. (N. Y.) 19, 14, Hinshw, L. B., M. M. Jordn, nd J. A. Vick. Histmine relese nd endotoxin shock in the primte. J. clin. Invest. 191, 4, Hlberg, F., nd W. W. Spink. The influence of Brucell somtic ntigen (endotoxin) upon the temperture rhythm of intct mice. Lb. Invest. 195, 5, Kbt, E. A., nd M. M. Myer. Experimentl Immunochemistry, 2nd ed. Springfield, Ill. Chrles C Thoms, 191, p Noh, J. W., nd A. Brnd. A fluorometric method to determine levels of histmine in humn plsm. J. Allergy 191, 32, Shore, P. A., A. Burkhlter, nd V. H. Cohn, Jr. A method for the fluorometric ssy of histmine in tissues. J. Phrmcol. exp. Ther. 1959, 127, Spink, W. W., nd J. Vick. Endotoxin shock nd the cogultion mechnism: modifiction of shock with epsilon-minocproic cid. Proc. Soc. exp. Biol. (N. Y.) 191, 1, Rmos, A. O., L. F. Chpmn, A. P. Corrdo, nd V. A. Fortes. Sympthomimetic ction of the epsilon-mino cproic cid. Arch. int. Phrmcodyn. 191, 132, Spink, W. W., nd J. Vick. Evlution of plsm, metrminol, nd hydrocortisone in experimentl endotoxin shock. Circult. Res. 191, 9, Kostk, J., nd J. Sterzl. The ction of endotoxin on complement. Foli microbiol. (Prh) 192, 7, Frnke, F. E. Attempts to produce nphylxis in guine pigs with polyscchride from Serrti mrcescens (Bcillus prodigiosus). U. S. Nt. Cncer Inst. J. 1944, 5, Wlkes, T. P., H. Weissbch, J. Bozicevich, nd S. Udenfriend. Serotonin nd histmine relese during nphylxis in the rbbit. J. clin. Invest. 1957, 3, Essex, H. E., nd W. D. Thoms. The response of the heptic venous circultion to certin substnces given intrvenously. Proc. stff Meeting Myo inic 195, 25, Gilbert, R. P., L. B. Hinshw, H. Kuid, nd M. B. Visscher. Effects of histmine, 5 hydroxytryptmine nd epinephrine on pulmonry hemodynmics with prticulr reference to rteril nd venous segment resistnces. Amer. J. Physiol. 1958, 194, Hddy, F. J. Effect of histmine on smll nd lrge vessel pressures in the dog foreleg. Amer. J. Physiol. 19, 198, Kuid, H., L. B. Hinshw, R. P. Gilbert, nd M. B. Visscher. The effect of grm-negtive endotoxin on the pulmonry circultion. Amer. J. Physiol. 1958, 192, Kowlewski, K., nd C. Wisniewski. Oxygen content nd ph of rteril, venous nd portl blood in dogs fter histmine. Proc. Soc. exp. Biol. (N. Y.) 192, 19, Schyer, R. W. Evidence tht induced histmine is n intrinsic regultor of the microregultory system. Amer. J. Physiol. 192, 22,. 27. Schyer, R. W. Induced synthesis of histmine, microcircultory regultion nd the mechnism of ction of the drenl glucocorticoid hormones in Progress in Allergy. Bsel, Switzerlnd, S. Krger, 193, vol. 7, p Wton, N. G. Is tissue histmine formtion necessry in ct, dog nd mn? Int. Arch. Allergy 193, 22, Crey, F. J., A. I. Brude, nd M. Zlesky. Studies with rdioctive endotoxin. III. The effect of tolernce on the distribution of rdioctivity fter intrvenous injection of Escherichi coli endotoxin lbeled with Cr51. J. clin. Invest. 1958, 37, Fine, J., H. Rutenberg, nd F. B. Schweinberg. The role of the reticulo-endothelil system in hemorrhgic shock. J. exp. Med. 1959, 11, Fox, C. L., Jr., nd S. E. Lsker. Protection by histmine nd metbolites in nphylxis, sclds, nd endotoxin shock. Amer. J. Physiol. 192, 22, Melby, J. C., R. H. Egdhl, I. C. Bossenmier, nd W. W. Spink. Suppression by cortisol of incresed serum-trnsminse induced by endotoxin. Lncet 1959, 1, Weissmnn, G., nd L. Thoms. Studies on lysosomes. I. The effects of endotoxin, endotoxin tolernce, nd cortisone on the relese of cid hydrolses from grnulr frction of rbbit liver. J. exp. Med , Spink, W. W. Endotoxin shock. Ann. intern. Med. 192, 57, 538.