- Email: [email protected]
Pathology of the active Arthus reaction in the chicken
Research in
cience 1983. :
ratnology of the active Arthus reaction in the chicken B. S. DHODAPKA,R, J. L. VEGAD, Gi. N. KOL'TE,Deparfment of 1)athology, College o,f Veterinai Science and Animal IHusbandty, Jabalp ur-482001, India, R. G. DHAVYEDKAR,,Departm,ent of Microbio'logy, Colhege of Vetc~rinatyScience and kl n imal Huabandty, labalpur-4'82001,In(fia
.
-~. .~
It was possible to produce anI active Atthus reaction in chicken skin which resultea In gross and microscopic lesions. Histologically, the reaction was p dominantly thrombotic in nature and restricted the upper dermis. The thrombi appeared to deve as a consequence of immune complex deposit with adherence and aggregation of thrombocytesi at the vascular endothelium. Thrombosis induced wi despread necrosis and haemorrhage and vascullitis occurred in the lower dermis. Up to four hours al inoculation, the cell population comprised infiltration of heterophils, monocytes and basoph suggesting an immediate hypersensitivity reactit This was followed by an Arthus type reaction for four to 12 hours involving both heterophils r~ n d monocytes. A characteristic feature was the devell ment of early perivascular lymphoid foci. After hours the reaction resembled a delayed hypersensitivity. The use of colloidal carbon suggested that whereas phagocytic activity of the heterophils and basophils appeared sensitisation dependent, that of thrombocytes and monocytes was independent oi The findings indicate that in the Arthus reaction the chicken the thrombocyte appears to be principal cell producing tissue damage by thrc,.-bosis. A (:ompariso~n was mad e with the active Artlhus reaction in the rabbit. -
-~
."LL
P
sequential gross and histopathlological changes. ... mat .. typically induced Comparison is also made wicn ie rabbit. 8
l.lPl
Ani,mak
Hiealthy White Leghorn vullets, 14 to 20 weeks
A
and healr:hy female rabbits, 2 weeks c)Id, were 1.
Bovine sertlrn albumin Cohn fraction V (BSA) ntron Rese:arch Laboratories, Bombay) was used. EXPbKIMLNl* L PROCEDlJRE -_2---. ~h~ design,of the two cxperlmenrs for stuciying the . . . * a * reaCtiG,r,a .vlGaGl,ted in Tapla
." ."
sitisation ( .
.
kperiment A. Four chickens and two rabbits were use(j. These were sensitised with a 5 per cent saline SOILtio on of BSA, repeated at four day intervals. In the chicken, the first injection of 0.2 ml was given . . " --.-.--ll. ,..4--..-:l.-, 4 -" I lnrravenously into the wing vein followed by five the haemorrhagic reaction developing in the skin c) f a intraperitoneal injections in doses of 0.2, 0.5, 0.5, sensitised animal after local injection of antig;en. 1 . 0 and 1 . 0 ml, respectively. The rabbits received More recently, Cochrane and Janoff (1974) sug- 0.5 , 0.5, 1.0, 1.0, 2.0 and 2 - 0 ml of the solution gested that it involved the deposition of antiboay- lntravenously into the ear vein, also at four day antigen complexes in the walls of Iressefs wh~ich inte activated complement and resulted in accumulation of neutrophils and subsequent tissue d anage. ~ Experrmenr s.cignt cnlcKens were sensirisea oy The Arthus reaction has served as a model of ac:Ute the method of Benedict (1967). The chickens were immunological injury from which a great deal of injemcted twice intravenously with a saline solution of information has been derived. However, rrlost BSL5. (40 and 20 mg/kg bodyweight, respectively) information on its pathology has been obtained fr,om . . wit1 . I a 15 day interval between injections. After seven studies in laboratory animals, mainly in the rabbit days, four sequential injections of BSA were given (Cochrane and Janoff 1974, Turk and Willoughby intramuscularly, at four day intervals, at a dose rate 1978). Virtually no information is available in the of 20 mg/kg bodyweight in normal saline mixed with chicken. This paper reports on the induction of the an equal amount of Freund's complete adjuvant active or classical A,rthus reaction in chickens and u
:-A-
i
terials and methods
B. S. Dhodapkar, J. L. Vegad, G. N. Kolte, R. G. Dhawedkar
260
TABLE 1: Experimental design for active Arthus reaction i n the skin ixperiment
Number of animals
Induction of the reaction
ne intervals lowing induc
Each sensitisled chicken received 13 d injections ...AL .,.&. ..-a rnoraclc --.. : . on errner lareral region of 0.2 ml saline solution containing 10 BSA. The injections were at 0.8.20.32. 44.48. 50, 52.53. 54.5.55 and 55.5 1 The birds were exsanguin at 56 h. This yielded lesio~ desired intervals
irteen sequel ,ions of 30 mi 1.5,2,3,4, EI, 12, 24, 36, 4i d 56 h duraticI n !re obtained in ch bird. They'were . . amined grossly and croscopically
-
4 chickens
ij
-L
2 rabbits
Sensitised chickens were given 0.2 ml c~fsaline id :----' , . I
----. .
(
As above. The lesions we mainly produced in the abdominal region ur lesions of 4 h d four lesions of 12 h were obtained from separate blirds. Four birds wereI used for each time interval. The lesions wer e , .-A ex,amined gross;iv ano microscopically
Sensitisationwas done bj the method of Benedict (1967). Each sensitised chicken received two id injections onI either lateral thorac:ic region of 0.2 ml saline solution containing 110 mg BSA
Sensitised chickens were given 0.2 ml saline id at correspondingintervals
id lntradermal
9f Arthus ireaction
.
"
.. . . .
.
--
I.-?--.
,La
'-
~- -
.~~.
..g. The erythema, oedema, necrosis and sloughin; n-..,. nnq r--. ut t i q ~ ~were l ~ s fixea m Larnoy s I"I U I ~ ,ana secn-..------thick. Sections were stained with haematoxylin 5~rm ande!osin, 0.05 per cent solution of toluidine blue in acetate buffer (pH 3.8 for 72 hours) for basophils dapkar et a1 1982), Mallory's phosphot---0':~ 1 1 5 ~ haemat ox]din for collagen, fiblrin and m uscular tures (Cullling 1974),, and 0.1 per cent a.queous nin. 3
u n e aay aetore tne inauctlon or m n u s react101n, feathers from the lateral thoracic regions we1re plucked and the skin cleaned using a soft clotLh moistened with sterile distilled water. Cleaning tl'e kin was repeated the following day. In rabbits, tl lair was clipped in the abdominal and thorac egion and the skin similarly cleaned. The acti~ Whus reaction (AAR) was induced by intraderm noculatiorI of antigen using a ieedle fitte )n a 1 ml t~lberculin syringe. ervations of the AP LR lesions differed i The obs~ L. ..- - . AAK . A m two experiments. In the flrst, lesions of tne larying duration (Table 1) were induced in the san mimal. In the second, to exclude the possibility ( nodification of the reactions occumng in the san mimal, four and 12 hour lesions were studie eparately im different groups of chickens. A colloidal uspension of carbon (Gunther Wagner Pelikai Nerke, bzitch C 11/1431 a) was injected intria. -, ... the wing vein at a dose rate of 0 - 1 m,, lenously Into 30 mir 00 g bc ore killir Awadhiya ).
-3
.
A ,,,
Evaluation of microscopic changes Extent and intensity were expressed on an arbitrary o f O , + , +, ++, + + + and + + + .t . The :ipation of leucocytes was determined by' countiem in five representative areas of vasclulitis in and medium venules. The number o'btained . was multiplied by the total number of lesions examined for each time interval (Dhodapkar et a1 1982)I.The counts presented in the tables represent meanI values with standard error. Thus, the cells were --. ..ted: intravascularly, ie, within the entire venule; rnurally, ie, covering the entire vessel wa 11; perivascu~larly,ie, in the immediate vicinity of thle entire .rnn..l le; diffusely in a high power microscop, GnlA Q), ie, all t he cells colvered und er this ma;gnifica-
.
# ;.,
Ciross ana r ll0gy All animals were k illed by exsanguination. Lesiol were excise:d and exaunined grc)ssly. Each lesion wi ---I -- an -- -.tirbitrarv scale of 0. I neasured, m u scvrtv VII t + + and + + respect ( ---A-
,,L,U
generate but recognisable cells wer,e also ..> --,I-A-L --> Leu. nowever cells In 1L~ n enecrotic ueorls anu ..-. ascular lynnphoid agl ere excluded. 7 ,
~
1-
2-
~ 1 ~
Arthus fieacnon m cnrctcens onv{ards the (:entres bec: m e necrlosed and (:yanotic. By! . . 56 hours, . . .rlecrosis ha d spread tc3 the periphery and
Results bXk'bKIMkN 1 A : ACI-IVEAK'IWUS KEALTlUN IN THE
CHICKEN AND RABBIT
Gross chc
-
Mic
01 AAK m cnlclten and rabbit were i, and are presented in Table 2.
Chicken. The microscopic changes of AAK in the chicken are presented in Table 3. At 30 minutes after inoculation, the changes comprised hyperaemia, mild oedema and haemorrhage around venules. Between one and four hours venules in the upper dermis showed fibrinoid necrosis. Small and medium ven ules showed thrombosis (Fig 2) with clumped thrc~mbocytes,fibrin and hyaline material. In the nation of venules of the lower dermis there was margi~ heterophils and of monocytes. These klad also infiltrated the vessel wall, which contained . . at. amorphous material. These changes were maximal two hours for heterophils and four hours for the molnocytes. Many emigrating cells contained phagocytc)sed amorphous material. The heterophils showed Aam,enerative changes at four hours. Basophils had I migrate howed dl on with
lesions
-. .
!n. I ne lesions exarnlnea up to ILnours were characterised by erythema and oedema. They urere well demarcated, oval and uniformly erythemat ous (Fig 1). Their intensity was maximum at two hoi1rs. The lesions from three hours onwards were less demarcated. After six hours Icentres shc)wed necnXIS and sloughing. Aftc:r 24 hours;,the erythlema was seen only peripherally. 1The 24, 36,, 4 8 and 5t5 hour lesions were necrotic and c:vanotic. Rabbit. Lesions examined up to 12 nours a'fter ---..1-a:. 3n were erythematoc1s and oedematous. 'rhe erythema was maximum between four and 12 ha,..ve although the cent]res were paler. Frc :
HlUFUlaLll
..
*
-.
...
TABLE 2: Cornpanson or gross lemons or scnve ~ r m u reacrlon s In m e c n ~ c ~ e ana n
Duratic or reac Lonrro
Animals
Siz (cr
Erythema
Necrosis and sloughing
Remarks
0.2 IT saline
Chicken (C) Rabbit (RJ
30 min
L'U
2.2 2.2 2.1 7.9
.2.1 1.a 2.0 1 .a
-
'Mean of four values of maximum diameter in chi1ckens and two in rabbits
L ~ L
B. 3. UnoaapKar, J . L. vegaa, CI. N .
-
-
Kolre, K .
CI.UnaweaKar
"i
<-
* - rdc
A
FIG 1 : Active Arthus reaction IAAR) on right lateral thoracic region of a chicken. Note oval, well demarcated and uniformly erythematous lesions of oneand a half, two, threeand four hours (left to right). The lesionsof 30 minutes and one hcJur are not y~n well develc
FIG 2: T nto-hour AAR . Section of Ichicken skin showing early thrombosis in a small vc?nule In the dermis (ri(9ht). and vaslculitis in the Iower dermis (left) H b E : 600
L
BLE 3: Microscopic chanaes in acthre Arthuo reamon m cnic:ken skin
u -u
T i interval
Hyper-
aemia
Oedema
30 min
+
+
-
rn-
I
--
rosisof vessel
iv lntravasculal im Intramural PV Perivascular . . >Cant, lsolat t Most of the cells necrotic Most of the cells degenerate but recognisable
*
*
-
Perivascul lympt , . ; , I "8"
fhlm bce
aggre gate!
anem Hetera
flit cells
-
Basophils
D. 3. UnoaapKar, J . L. vegaa, ti. N .
LW
norre, K .
ti. unaweaKar
*m
I*
FIG 3: Eight-hour AAR. Sect~onof chlcken skin show~ngf~brlno~d necrosls of venular wall and preser hyallne material in the lumen. H 8 E x 600
maxlmum intensity at three hours. Some of the ?enules in the deeper dermi!r showed mild per,i,ascular lymphoid aggregations The six- and eight-hour le:sions we1-e intense:ly -L -1 Lf- r.,f laemorrhalzic and necrotic andI srruweu sluugnrrlg epidermis in places,. The venule walls showing fibrinoid Inecrosis \were thickened and intensely eosinophilic and . . con1 .-. tained thrombocytes, fibrin and hyaline material (Pig 3). Some of the small venule~ vere completely occluded by hyaline thrombi (Fig 4) -eucocyte emigration was marked and perivascul;u ymphoid aggregations had increased. In the deepc:r lermis, muscle fibres showed hvalinisation and foc,"1 dl nyositis. The lesicDns from 12 hours, onwards were prcI.ressively Iless . oeden .. latous antd. .more necrotic . . an~d ~aemorrhagic.kpiaerma sloughing was wdespreac1. it 12 hours the perivascular lymphoid aggregatiorIS tere two to four cells thick. Between 24 to 48 hour s, he aggregations had spread resulting in confluenc:e df neighbouring foci (Fig 5). They also revealed a few blast cells. Occasionally, vessels contained hyalirhe thrombi predominating in lymphocytes (Fig 6). Ttle subepidermal necrotic tissue showed marke,d ccumulatiion of degcmerate heterophils. The monc ytic cells were vacu olated, sc)me presel~ t i n gsigncDt ing appealmce. Epithelioid ceblls, macrophages an d . . cells surroundea,rne necrotic tissue. lccasional gianr The thrombi becarne progre:ssively more compac:t lrith increased hyaline material (Fig 7). A~t48 hour:s -0
-
..
a.
.a
the thrombi in the upper dermis completely occluded even the larger venules (I:ig 8). Tht:y revealeci either wides pread nelcrosis or massive haemorrh age in surrounding tis:rue. -'L - --:2--At 56 hours rrrusr ur rue euruermis had sl,ounhed awav. Perivasc:ular lyml3hoid aggregations in the lower dermis wc:re pronou nced. A..--
A
Rnbblt. I he microscopic changes ot AAK *n the rabbi t are presented in Table 4. At 30 minultes the changges comprised hyperaemia and mild o edema. After one hour there was progressive emigration of I-..-mainly neutro~hils. Other c.I-----I C U C Ucytes, ~ ;rlarlgcs inclucied necrosis of the ressh wall and thrornbosis. Throimbi contained plate1ets, neutrc)phils and fibrin. Oede: . "ma and haemorrhagc:were moc .. ierate. Afl er 12 hours the neutropnlls were degenerate, maki~ 1g mononuclear infiltration more consp icuous. Eosin ophils were also seen, particularly around necrotic tissue and thrombi were more compact. At 56 hcJurs, ther e was slc f epiderm widespread necl.osis. a.a . .
.A
L..b
.
tXt'tI
- .. ... iIMhN 1 L): AL I L V t A K L H U S K t A C T l U N I N .-
CHlCE
FourGri ., lly spherical leslonC < w i t h -. .3ss cnang--. moderate eryth ema and oedema nneasured : 0.041:m.
Arthus reaction in chickens
FIG 4: Eight-hour AAR. Section of chicken skin showing a venule (left) completely occluded by a hyaline thrombus Venule on the right shows early stages of thrombosis. H 8 E x 300
;.
FIG 5: 'rwenty-four-hour AAR. aggregates. H 6 E x 600
E
icken skin stlowing conflluence ot nei!
nphoid
266
B. S. Dhodapkar, J. L. Vegad, G. N. Kolte, R. G. Dhawedkar
FIG 6: Thirty-six-hour AAR. Section of chicken skin showing a vein with a hyalinethrombus predominating ded by a lymlphoid aggregation. H & E in lymphocytes. The vc
FIG 7: Twtenty-four-hoiur AAR. Section of chicken skin showring thrombo~ with necn3sis of its wa II and the intctrstitium. H Et E x 3 0 0
thrombus in i9 vein
TABLE 4: Microscopic changes in active Arthus reaction in tab~ bskin i
-
Neutrophils
Monocyticcells
Diffuse
0n
t t
-
12 h
T
t
++
lntravasculi Intramural r Perivascula~ Scant, isolalted cells .a-
-A
t
-'-L.
Most of the cells degene
f
t
t
T
++
t
Eosinophils
Diffuse
4. S.
Dhodapkar, J. L. Vegad, G. N. Kolte, R. G. Dhawedkar Microscopic changes. These are presented in Table 5. The changes in the upper dermis comprised hyperaemia, oedema, haemorrhage, fib]Finoid necrosis and thrombosis. Phagocytosed carbon was seen in thrombocyte aggregates. The changes in the lower dermis included vasculitis, perivasculitis: with marked heterophil infiltration and necrosis of the vessel wall. Some venules revealed carbon labelling, while others showed marked spread of carbon into the adjacent tissues. Almost all the heterophils and monocytic cells, both in and outs;ide the vessels contained engulfed carbonI and amorphous material and mostly (Fig 9). Basophils showed degranula . . tion . I here were )&id aggregations. moder,ate perivas
...,..
Twelve-hourAA Gro.s changes. Roughly spherical lesions with moder ate erythe:ma and extensive oedema measured 2.93 ?: 0.04 cm. Necrosis and sloughing of epidermis ,.. :Aa.., were a I",. l~u svlusrll Miciroscopic Ichanges. These arle present'ed in Table 5 . Chang es in the upper. dermis comprised extensive oeaema, tnromDosis, necrosls and haemorrhage. The lower dermis revealed moderate lymphoid aggregations with numerous blast cells. The predominant cells were monocytic and contained a varyiing propor ,tion of ca:rbon. The Arthus reaction in the c:hicken an d rabbit is comparedI in Table (
.
Discussion The present stuay nas confirmed the earlier report of Luoma and Benedict (1977) that it is possi ble to elicit an Arthus reaction in the chicken. HolNever, much higher doses of innmune re;actants may be required to obtain reactioris of opticnum inten sity in older birds of 14 to 20 weel
Arrnus reacrron zn cnrcKens
FIG 8: Forty-eight-hour AAR. Section of chick 'hrombus. H & E x 600
ving a large \
h-1;"-
ur-hourAAR. Section of chicken skin showing heterophilsand a few monocytic cells, inandaround vith engulfed
TABLE 6: Comparison of active Arthus reaction in the chicken and rabbit
In 60 min Less inten )r~rtc~n ur~uuaa n ru vcucmr~atvua.
Intensity
A..,.
,.&.-.-
atous
atnu v r u = ~ ~ r r ~ v u LS a ".u 8 . 3 ,
,a
so covered a wide area
restricted to the periph~ ew, particularly between 4 iand 12 h
aximal betwea n 2 a n d 6 h
Maximal between 4 ancJ 8 h
,.,.,.,
,., ..,.
2nd nnt
ex
Commence from 24 h, extensive
Predominantly thrombotic; less inflamrnatc
Predominimtly inflamm less throrr.L-*:-
Hc?terophilsfor1med only a par t of the inflalmmatory re;action
lnflammatory reaction predominimtly neutrophilic from 1 to IB h
Monocyte!;(monocytic cellsf large mononuclea cells and niacrophages)
Participation of almost similar intensity right from the early stages
Little participation before 6 to 8 h
Lymhocytc (large and
Slight to moderate, predominantly perivascular lymphoid aggrergation from 3 to 8 h, and intense aggregation frcIm 12 h onwards
Only rarely in aggregates; more diffuIse and appe; at a much later stage (is from 12 h onwards)
Necrosis and icroscopic ch Vascular cha
-
m ,.
.mmnnrs 2 - . +
...
6 h, and
BaSOpnllS
gnificant pad:clpanon 2to4h
No significanr panlctpanon at 2 to 4 h
Eosinophil:
3t definite; d ifficult to sess
Significant in numbers at 24 to 48 h
lent of perivascular lymphoid aggregaIns; and 4, resemblance of AAR wit h delayedI persensitivity in the later stage!S. The most prominent feature of the re: iction was; '- -. e formation of thrombi in the early stages. a".~ n- i s is preceded by hyaline degeneration of the vessel ills. While the precise nature of the hyaline aterial remains unknown, it could be a conse.dence of antigen-antibody reaction (Jonsson 1972). In the chicken, thrombi were confined to the upper dermis away from areas of inflammation. It is possible that higher concentrations of immune re;actants in this region may attra.ct thrombocytes and in:itiate thrombosis. In the rabbit, thrombosis has be,en attributed to glycolytic prcoducts of leucocytes - - -. - - endothelium -. wllich are toxic to vascular (Stetson 19151), or to the damage effected by vascular inflam.. m;ation (Cochrane and Janoff 1974). The presenl: re:sults indicate that in the chicken thrombosis does; n o)t appear to be a consequence of inflammatory .scular dannage. The fibrin negative amorpkIOUS mate:rials seenI e in the lulmen of the: vessel are ling the wi311 and fre~ -..- - currlutelrrs 1 ,nL->..- .1.-~parently .immunt: turluuaorrar e-. l a1 ,cur in the 182). Similar deposit -
- -
-~~~
-1
-I
..,
reaction la^^^^. I firbar, l r a u r r a JU~;~;GJL that th rombus formation in the upper dermis begins with dc:position of immune complexes, adherence of throml3ocytes to the injured endothelium and their -. aggregation. It is also likely that the chicken thrombocyte may be engulfing immune complexes since it is established that it is a phagocytic cell (Glick et a1 1964, Carlson et a1 1968, Chang and Hamilton 1979, Awadhiya et a1 1980). The present stud:Y also revealed thrombocytes with engulfed carbon. Recently, in rabbit basophils, the presence of a platelet aggregating factor has been demons trated (Frick 1980). While nothing is known in the chicken, it may be significant that basophils were seen participating in the early Arthus reaction in considerable numbers and mav have a role in thrombocvte aggreg;ation. The inflammatory chanlges in th e chicken were confinced to the 1ower dernnis. -This nnight have: been . due to a low concentration of immune reactants in this region. As in the rabbit, the immune reactants may activate complement and produce factors chemotactic to leucocytes (Cochrane and Janoff N C,.U$
---
-
1974).
he chicke,n, the in~itial emig
comprised heterophils. This was soon followed by ing with various stimuli they reported the aggregates concurrent emigration of monocytic cells and occurring much later (Carlson and Allen 1970, basophils. A similar mixed exudate has been report ed Carlson 1972, Nair 1973, Awadhiya et al 1981a). by Chand et al (1976) in passive cut;aneous anla- Earl:y developnnent of lynnphoid aggregations in AAR phylaxis. Mixed cellular responses havse even been suggests it to ble a respon!;e to the antigenic stimulus. n _-.-._.: ~. ns Tt reported in non-immunoloeical inr~ammarlo --- -le lesions at ~4 and 48 hours histoloeicallv - u-----.-=----a (Carlson 1972, Nair 1973, A.wadhiya Iet a1 1981a, resernbled delayed hypersensitivity reactions (Ander1981b). The present results in1dicate thait the inflam- son 1971, Cheville and Richards 1971, War1ier et al matory changes in the chicker1 were lessI severe th an 1971, Rose and Bradley 1977). Despite the a pparent those in the rabbit. In tl.,h a latter neutrophils resemblance, AAR cannot be considered a!r3 oa +Li-..a dominated the early reaction (Gel1 and Hinde 195i4, dela!led hypersensitivity reaction iin the late1. stages, Humphrey 1955, Cochrane and Weigle 19513), asit is initiated by antigc:n-antiboc ly comple~ ces, and whereas in the sheep, there was delayed migration of its a:ssociation with cellurlar immu;nity has not been >.-. neutrophils and an early infiltration of mononuclear aernonstrated. cells (Vegad and Lancaster 1972, Vegad 1979). Bas on this work it appears that heterophils:,monocyl and basophils are involved in early inflixmmation 7 cllfiallthe chicken. Heterol:jhils were actively piiagocytic. In the fotlr- Werlke, Hanover, West Germany for the su~ P P ~ofY ~ O carbon. Our thanks are also due to I)r R. P. hour AAIR -in the by repeat ed C O ~ lidal . chicken, riensitised .Awa dhiya and the dean, Dr B. S. Malik f or their injections of antlgen in Freund's complete adjuva~ It, and to Dr A. G. Khan, senior scientis. almost all the heterophils in the exudate revealed Interest D,...l try Project, for the supply of b irds. phagocytosed carbon and the amorphous materi; Awadhiya et a1 (1980) demonstrated a lack of phag Recelived for pr ublication December cytic activitv . bv. hetero~hilsin earlv inflammatia Hence thle presenlt inding!r suggest that thleir phagocytic: propert ies may depend upon thleir References sensitisaticIn. The early appearance ot monocytes suggests their ANDERSON. J. C. (1971) Research in Veterinary Sc;nn- 1' 312-317 possible role in phagocytosis and degradation of R. P., VEGACI, J. L. & ULOLTE. G. immune complexes, thus supporting heterophils in AWADHIYA, Research in Veterenary Sciencc?29,120-122 the process. The monocytes appeared more uniform AWADHIYA, R. P.. VEGAD, I. L. & KiOLTE, G. b ".:.~.. Research in Vetennary acrence 31.23 1-235 in their phagocytic activity towards carbon and AWADHIYA, R. P.. VEGAD ULIE, cr. n. (IYUID) immune complexes, regardless of sensitisation. These Avian Pathology 10,3 13-320 findings are in contrast to those in the rabbit, in BENEDICT. A. A. (1%7) Methoas m immunology and Immunowhich the antigen is revealed by neutrophils in the chemistry, Vol 1. Eds C. A. Williams and M. W. Chase. New York, Academic Press, pp 229-234 early hours, but not by mononuclear cells (Cochrane et a1 1959). Further studies arc: obviousl! required to CARLSON, H. C. (1972)AvionDiseuses 16. ! H. C. & ALLEN, J. R. (1978 elucidate the functional signiificance o f monocytic CARLSON, 817-833 cells in avian inflammation. CARLSON, H. C., SWEENY, P. R. & TAKARYK, J. in Diseases 12,700-715 The basophils were not only seen intra- and extl 4D. N., CARLSON, H. (::. & EYRE, :P. (1976) Int~ vascularly, but were also found phagocytising carbl Arc hives of Allergy and Appli,ed Immunolorgy51,508-5117 particles. Dhodapkar et a1 (1982) described phagloCHAP4G, C. F . & HAMILT(IN, P. B. (:1979) Journa1 of the cytosis of carbon by basophils in reverse (passijre) P .~cuIoendothelial ~, -R.-. Society 25.-585-5'30 - -- - - Arthus reaction. This property of basoptlils has again CHEVILLE, N. F. & RICHARDS, W. D. (1971)Americom Journal of P a ~ h o l o g64,97-114 ~ been demonstrated. The present filndings thus COCHRANE. C. G. & JANOFF. A. (1974) The Infl endorse the possibility that tmsophils Inay share a Process, Vol 2. Eds B. W. Zweifach, L. Grant a1 phagocytic property with other cells under immuno.'"-'Jluskey. New York. Academic Press. py,OJ-I"L 68) Journal I logic stim~ 1lus. COCFIRANE. C. G. & WEIGL menrtolMedicine 108,591 404 Anothelr noteworthy feature of Ithe chicken DIXON. F. COCFIRANE. C. G., WEIGLI reaction 1was the formation of early perivasculIar -494 . . Jou rnol of Evperimenlal Medrctne nu, 481.. onnranot, P r n m 9 A hn.. rc n"..,"v .A* -... . lymphoid awjljlc5arca. I nvrrn UJ CULLING, C. F. A. (1974) Handbook of Histopatnological Techaggregates became more cornpact an(1 tended to niques. 3rd edn. London, Buttenvorths. p i N. (1982) coalesce. There was a prepon derance of monocytic DHODAPKAR, B. S., VEGAD, J. L. & 1 Research in VeterinaryScience 33,377-379 cells among cellular infiltrations. The ccurrence of FRICK, 0 . L. (1980) Basic and Clinical Immunology. EIds H. H. .> .-.~....--. 1s consistent - -- -2 ..: perivascular lymphoia aggregations wNn Fud enberg, D. P . Stites, J . L. Caldwell andI J. V. Wells. Maruzea. Lange Medical Publications. r,p 274-294 the findings of other investigaltors. Hovvever, WOI'k- 3
.,.,.
LA..
.I
* A .
! . A .
-2
e .
I",L\
or
-
AlWU L D
Wll"41
08
?.
I
L
L
-- - .
OC
lL3
,.
LOUGHBY. D. A. (1978:I InflammaticJn. Eds ;. H. Ferreura. New Yo1ik, Springer-Verlag.
I r
VEGAD VEGAD Veteri, WARNE notion
. . ..
k KANTOR,
) Inter-
nv 40.
cience 1983. :
ratnology of the active Arthus reaction in the chicken B. S. DHODAPKA,R, J. L. VEGAD, Gi. N. KOL'TE,Deparfment of 1)athology, College o,f Veterinai Science and Animal IHusbandty, Jabalp ur-482001, India, R. G. DHAVYEDKAR,,Departm,ent of Microbio'logy, Colhege of Vetc~rinatyScience and kl n imal Huabandty, labalpur-4'82001,In(fia
.
-~. .~
It was possible to produce anI active Atthus reaction in chicken skin which resultea In gross and microscopic lesions. Histologically, the reaction was p dominantly thrombotic in nature and restricted the upper dermis. The thrombi appeared to deve as a consequence of immune complex deposit with adherence and aggregation of thrombocytesi at the vascular endothelium. Thrombosis induced wi despread necrosis and haemorrhage and vascullitis occurred in the lower dermis. Up to four hours al inoculation, the cell population comprised infiltration of heterophils, monocytes and basoph suggesting an immediate hypersensitivity reactit This was followed by an Arthus type reaction for four to 12 hours involving both heterophils r~ n d monocytes. A characteristic feature was the devell ment of early perivascular lymphoid foci. After hours the reaction resembled a delayed hypersensitivity. The use of colloidal carbon suggested that whereas phagocytic activity of the heterophils and basophils appeared sensitisation dependent, that of thrombocytes and monocytes was independent oi The findings indicate that in the Arthus reaction the chicken the thrombocyte appears to be principal cell producing tissue damage by thrc,.-bosis. A (:ompariso~n was mad e with the active Artlhus reaction in the rabbit. -
-~
."LL
P
sequential gross and histopathlological changes. ... mat .. typically induced Comparison is also made wicn ie rabbit. 8
l.lPl
Ani,mak
Hiealthy White Leghorn vullets, 14 to 20 weeks
A
and healr:hy female rabbits, 2 weeks c)Id, were 1.
Bovine sertlrn albumin Cohn fraction V (BSA) ntron Rese:arch Laboratories, Bombay) was used. EXPbKIMLNl* L PROCEDlJRE -_2---. ~h~ design,of the two cxperlmenrs for stuciying the . . . * a * reaCtiG,r,a .vlGaGl,ted in Tapla
." ."
sitisation ( .
.
kperiment A. Four chickens and two rabbits were use(j. These were sensitised with a 5 per cent saline SOILtio on of BSA, repeated at four day intervals. In the chicken, the first injection of 0.2 ml was given . . " --.-.--ll. ,..4--..-:l.-, 4 -" I lnrravenously into the wing vein followed by five the haemorrhagic reaction developing in the skin c) f a intraperitoneal injections in doses of 0.2, 0.5, 0.5, sensitised animal after local injection of antig;en. 1 . 0 and 1 . 0 ml, respectively. The rabbits received More recently, Cochrane and Janoff (1974) sug- 0.5 , 0.5, 1.0, 1.0, 2.0 and 2 - 0 ml of the solution gested that it involved the deposition of antiboay- lntravenously into the ear vein, also at four day antigen complexes in the walls of Iressefs wh~ich inte activated complement and resulted in accumulation of neutrophils and subsequent tissue d anage. ~ Experrmenr s.cignt cnlcKens were sensirisea oy The Arthus reaction has served as a model of ac:Ute the method of Benedict (1967). The chickens were immunological injury from which a great deal of injemcted twice intravenously with a saline solution of information has been derived. However, rrlost BSL5. (40 and 20 mg/kg bodyweight, respectively) information on its pathology has been obtained fr,om . . wit1 . I a 15 day interval between injections. After seven studies in laboratory animals, mainly in the rabbit days, four sequential injections of BSA were given (Cochrane and Janoff 1974, Turk and Willoughby intramuscularly, at four day intervals, at a dose rate 1978). Virtually no information is available in the of 20 mg/kg bodyweight in normal saline mixed with chicken. This paper reports on the induction of the an equal amount of Freund's complete adjuvant active or classical A,rthus reaction in chickens and u
:-A-
i
terials and methods
B. S. Dhodapkar, J. L. Vegad, G. N. Kolte, R. G. Dhawedkar
260
TABLE 1: Experimental design for active Arthus reaction i n the skin ixperiment
Number of animals
Induction of the reaction
ne intervals lowing induc
Each sensitisled chicken received 13 d injections ...AL .,.&. ..-a rnoraclc --.. : . on errner lareral region of 0.2 ml saline solution containing 10 BSA. The injections were at 0.8.20.32. 44.48. 50, 52.53. 54.5.55 and 55.5 1 The birds were exsanguin at 56 h. This yielded lesio~ desired intervals
irteen sequel ,ions of 30 mi 1.5,2,3,4, EI, 12, 24, 36, 4i d 56 h duraticI n !re obtained in ch bird. They'were . . amined grossly and croscopically
-
4 chickens
ij
-L
2 rabbits
Sensitised chickens were given 0.2 ml c~fsaline id :----' , . I
----. .
(
As above. The lesions we mainly produced in the abdominal region ur lesions of 4 h d four lesions of 12 h were obtained from separate blirds. Four birds wereI used for each time interval. The lesions wer e , .-A ex,amined gross;iv ano microscopically
Sensitisationwas done bj the method of Benedict (1967). Each sensitised chicken received two id injections onI either lateral thorac:ic region of 0.2 ml saline solution containing 110 mg BSA
Sensitised chickens were given 0.2 ml saline id at correspondingintervals
id lntradermal
9f Arthus ireaction
.
"
.. . . .
.
--
I.-?--.
,La
'-
~- -
.~~.
..g. The erythema, oedema, necrosis and sloughin; n-..,. nnq r--. ut t i q ~ ~were l ~ s fixea m Larnoy s I"I U I ~ ,ana secn-..------thick. Sections were stained with haematoxylin 5~rm ande!osin, 0.05 per cent solution of toluidine blue in acetate buffer (pH 3.8 for 72 hours) for basophils dapkar et a1 1982), Mallory's phosphot---0':~ 1 1 5 ~ haemat ox]din for collagen, fiblrin and m uscular tures (Cullling 1974),, and 0.1 per cent a.queous nin. 3
u n e aay aetore tne inauctlon or m n u s react101n, feathers from the lateral thoracic regions we1re plucked and the skin cleaned using a soft clotLh moistened with sterile distilled water. Cleaning tl'e kin was repeated the following day. In rabbits, tl lair was clipped in the abdominal and thorac egion and the skin similarly cleaned. The acti~ Whus reaction (AAR) was induced by intraderm noculatiorI of antigen using a ieedle fitte )n a 1 ml t~lberculin syringe. ervations of the AP LR lesions differed i The obs~ L. ..- - . AAK . A m two experiments. In the flrst, lesions of tne larying duration (Table 1) were induced in the san mimal. In the second, to exclude the possibility ( nodification of the reactions occumng in the san mimal, four and 12 hour lesions were studie eparately im different groups of chickens. A colloidal uspension of carbon (Gunther Wagner Pelikai Nerke, bzitch C 11/1431 a) was injected intria. -, ... the wing vein at a dose rate of 0 - 1 m,, lenously Into 30 mir 00 g bc ore killir Awadhiya ).
-3
.
A ,,,
Evaluation of microscopic changes Extent and intensity were expressed on an arbitrary o f O , + , +, ++, + + + and + + + .t . The :ipation of leucocytes was determined by' countiem in five representative areas of vasclulitis in and medium venules. The number o'btained . was multiplied by the total number of lesions examined for each time interval (Dhodapkar et a1 1982)I.The counts presented in the tables represent meanI values with standard error. Thus, the cells were --. ..ted: intravascularly, ie, within the entire venule; rnurally, ie, covering the entire vessel wa 11; perivascu~larly,ie, in the immediate vicinity of thle entire .rnn..l le; diffusely in a high power microscop, GnlA Q), ie, all t he cells colvered und er this ma;gnifica-
.
# ;.,
Ciross ana r ll0gy All animals were k illed by exsanguination. Lesiol were excise:d and exaunined grc)ssly. Each lesion wi ---I -- an -- -.tirbitrarv scale of 0. I neasured, m u scvrtv VII t + + and + + respect ( ---A-
,,L,U
generate but recognisable cells wer,e also ..> --,I-A-L --> Leu. nowever cells In 1L~ n enecrotic ueorls anu ..-. ascular lynnphoid agl ere excluded. 7 ,
~
1-
2-
~ 1 ~
Arthus fieacnon m cnrctcens onv{ards the (:entres bec: m e necrlosed and (:yanotic. By! . . 56 hours, . . .rlecrosis ha d spread tc3 the periphery and
Results bXk'bKIMkN 1 A : ACI-IVEAK'IWUS KEALTlUN IN THE
CHICKEN AND RABBIT
Gross chc
-
Mic
01 AAK m cnlclten and rabbit were i, and are presented in Table 2.
Chicken. The microscopic changes of AAK in the chicken are presented in Table 3. At 30 minutes after inoculation, the changes comprised hyperaemia, mild oedema and haemorrhage around venules. Between one and four hours venules in the upper dermis showed fibrinoid necrosis. Small and medium ven ules showed thrombosis (Fig 2) with clumped thrc~mbocytes,fibrin and hyaline material. In the nation of venules of the lower dermis there was margi~ heterophils and of monocytes. These klad also infiltrated the vessel wall, which contained . . at. amorphous material. These changes were maximal two hours for heterophils and four hours for the molnocytes. Many emigrating cells contained phagocytc)sed amorphous material. The heterophils showed Aam,enerative changes at four hours. Basophils had I migrate howed dl on with
lesions
-. .
!n. I ne lesions exarnlnea up to ILnours were characterised by erythema and oedema. They urere well demarcated, oval and uniformly erythemat ous (Fig 1). Their intensity was maximum at two hoi1rs. The lesions from three hours onwards were less demarcated. After six hours Icentres shc)wed necnXIS and sloughing. Aftc:r 24 hours;,the erythlema was seen only peripherally. 1The 24, 36,, 4 8 and 5t5 hour lesions were necrotic and c:vanotic. Rabbit. Lesions examined up to 12 nours a'fter ---..1-a:. 3n were erythematoc1s and oedematous. 'rhe erythema was maximum between four and 12 ha,..ve although the cent]res were paler. Frc :
HlUFUlaLll
..
*
-.
...
TABLE 2: Cornpanson or gross lemons or scnve ~ r m u reacrlon s In m e c n ~ c ~ e ana n
Duratic or reac Lonrro
Animals
Siz (cr
Erythema
Necrosis and sloughing
Remarks
0.2 IT saline
Chicken (C) Rabbit (RJ
30 min
L'U
2.2 2.2 2.1 7.9
.2.1 1.a 2.0 1 .a
-
'Mean of four values of maximum diameter in chi1ckens and two in rabbits
L ~ L
B. 3. UnoaapKar, J . L. vegaa, CI. N .
-
-
Kolre, K .
CI.UnaweaKar
"i
<-
* - rdc
A
FIG 1 : Active Arthus reaction IAAR) on right lateral thoracic region of a chicken. Note oval, well demarcated and uniformly erythematous lesions of oneand a half, two, threeand four hours (left to right). The lesionsof 30 minutes and one hcJur are not y~n well develc
FIG 2: T nto-hour AAR . Section of Ichicken skin showing early thrombosis in a small vc?nule In the dermis (ri(9ht). and vaslculitis in the Iower dermis (left) H b E : 600
L
BLE 3: Microscopic chanaes in acthre Arthuo reamon m cnic:ken skin
u -u
T i interval
Hyper-
aemia
Oedema
30 min
+
+
-
rn-
I
--
rosisof vessel
iv lntravasculal im Intramural PV Perivascular . . >Cant, lsolat t Most of the cells necrotic Most of the cells degenerate but recognisable
*
*
-
Perivascul lympt , . ; , I "8"
fhlm bce
aggre gate!
anem Hetera
flit cells
-
Basophils
D. 3. UnoaapKar, J . L. vegaa, ti. N .
LW
norre, K .
ti. unaweaKar
*m
I*
FIG 3: Eight-hour AAR. Sect~onof chlcken skin show~ngf~brlno~d necrosls of venular wall and preser hyallne material in the lumen. H 8 E x 600
maxlmum intensity at three hours. Some of the ?enules in the deeper dermi!r showed mild per,i,ascular lymphoid aggregations The six- and eight-hour le:sions we1-e intense:ly -L -1 Lf- r.,f laemorrhalzic and necrotic andI srruweu sluugnrrlg epidermis in places,. The venule walls showing fibrinoid Inecrosis \were thickened and intensely eosinophilic and . . con1 .-. tained thrombocytes, fibrin and hyaline material (Pig 3). Some of the small venule~ vere completely occluded by hyaline thrombi (Fig 4) -eucocyte emigration was marked and perivascul;u ymphoid aggregations had increased. In the deepc:r lermis, muscle fibres showed hvalinisation and foc,"1 dl nyositis. The lesicDns from 12 hours, onwards were prcI.ressively Iless . oeden .. latous antd. .more necrotic . . an~d ~aemorrhagic.kpiaerma sloughing was wdespreac1. it 12 hours the perivascular lymphoid aggregatiorIS tere two to four cells thick. Between 24 to 48 hour s, he aggregations had spread resulting in confluenc:e df neighbouring foci (Fig 5). They also revealed a few blast cells. Occasionally, vessels contained hyalirhe thrombi predominating in lymphocytes (Fig 6). Ttle subepidermal necrotic tissue showed marke,d ccumulatiion of degcmerate heterophils. The monc ytic cells were vacu olated, sc)me presel~ t i n gsigncDt ing appealmce. Epithelioid ceblls, macrophages an d . . cells surroundea,rne necrotic tissue. lccasional gianr The thrombi becarne progre:ssively more compac:t lrith increased hyaline material (Fig 7). A~t48 hour:s -0
-
..
a.
.a
the thrombi in the upper dermis completely occluded even the larger venules (I:ig 8). Tht:y revealeci either wides pread nelcrosis or massive haemorrh age in surrounding tis:rue. -'L - --:2--At 56 hours rrrusr ur rue euruermis had sl,ounhed awav. Perivasc:ular lyml3hoid aggregations in the lower dermis wc:re pronou nced. A..--
A
Rnbblt. I he microscopic changes ot AAK *n the rabbi t are presented in Table 4. At 30 minultes the changges comprised hyperaemia and mild o edema. After one hour there was progressive emigration of I-..-mainly neutro~hils. Other c.I-----I C U C Ucytes, ~ ;rlarlgcs inclucied necrosis of the ressh wall and thrornbosis. Throimbi contained plate1ets, neutrc)phils and fibrin. Oede: . "ma and haemorrhagc:were moc .. ierate. Afl er 12 hours the neutropnlls were degenerate, maki~ 1g mononuclear infiltration more consp icuous. Eosin ophils were also seen, particularly around necrotic tissue and thrombi were more compact. At 56 hcJurs, ther e was slc f epiderm widespread necl.osis. a.a . .
.A
L..b
.
tXt'tI
- .. ... iIMhN 1 L): AL I L V t A K L H U S K t A C T l U N I N .-
CHlCE
FourGri ., lly spherical leslonC < w i t h -. .3ss cnang--. moderate eryth ema and oedema nneasured : 0.041:m.
Arthus reaction in chickens
FIG 4: Eight-hour AAR. Section of chicken skin showing a venule (left) completely occluded by a hyaline thrombus Venule on the right shows early stages of thrombosis. H 8 E x 300
;.
FIG 5: 'rwenty-four-hour AAR. aggregates. H 6 E x 600
E
icken skin stlowing conflluence ot nei!
nphoid
266
B. S. Dhodapkar, J. L. Vegad, G. N. Kolte, R. G. Dhawedkar
FIG 6: Thirty-six-hour AAR. Section of chicken skin showing a vein with a hyalinethrombus predominating ded by a lymlphoid aggregation. H & E in lymphocytes. The vc
FIG 7: Twtenty-four-hoiur AAR. Section of chicken skin showring thrombo~ with necn3sis of its wa II and the intctrstitium. H Et E x 3 0 0
thrombus in i9 vein
TABLE 4: Microscopic changes in active Arthus reaction in tab~ bskin i
-
Neutrophils
Monocyticcells
Diffuse
0n
t t
-
12 h
T
t
++
lntravasculi Intramural r Perivascula~ Scant, isolalted cells .a-
-A
t
-'-L.
Most of the cells degene
f
t
t
T
++
t
Eosinophils
Diffuse
4. S.
Dhodapkar, J. L. Vegad, G. N. Kolte, R. G. Dhawedkar Microscopic changes. These are presented in Table 5. The changes in the upper dermis comprised hyperaemia, oedema, haemorrhage, fib]Finoid necrosis and thrombosis. Phagocytosed carbon was seen in thrombocyte aggregates. The changes in the lower dermis included vasculitis, perivasculitis: with marked heterophil infiltration and necrosis of the vessel wall. Some venules revealed carbon labelling, while others showed marked spread of carbon into the adjacent tissues. Almost all the heterophils and monocytic cells, both in and outs;ide the vessels contained engulfed carbonI and amorphous material and mostly (Fig 9). Basophils showed degranula . . tion . I here were )&id aggregations. moder,ate perivas
...,..
Twelve-hourAA Gro.s changes. Roughly spherical lesions with moder ate erythe:ma and extensive oedema measured 2.93 ?: 0.04 cm. Necrosis and sloughing of epidermis ,.. :Aa.., were a I",. l~u svlusrll Miciroscopic Ichanges. These arle present'ed in Table 5 . Chang es in the upper. dermis comprised extensive oeaema, tnromDosis, necrosls and haemorrhage. The lower dermis revealed moderate lymphoid aggregations with numerous blast cells. The predominant cells were monocytic and contained a varyiing propor ,tion of ca:rbon. The Arthus reaction in the c:hicken an d rabbit is comparedI in Table (
.
Discussion The present stuay nas confirmed the earlier report of Luoma and Benedict (1977) that it is possi ble to elicit an Arthus reaction in the chicken. HolNever, much higher doses of innmune re;actants may be required to obtain reactioris of opticnum inten sity in older birds of 14 to 20 weel
Arrnus reacrron zn cnrcKens
FIG 8: Forty-eight-hour AAR. Section of chick 'hrombus. H & E x 600
ving a large \
h-1;"-
ur-hourAAR. Section of chicken skin showing heterophilsand a few monocytic cells, inandaround vith engulfed
TABLE 6: Comparison of active Arthus reaction in the chicken and rabbit
In 60 min Less inten )r~rtc~n ur~uuaa n ru vcucmr~atvua.
Intensity
A..,.
,.&.-.-
atous
atnu v r u = ~ ~ r r ~ v u LS a ".u 8 . 3 ,
,a
so covered a wide area
restricted to the periph~ ew, particularly between 4 iand 12 h
aximal betwea n 2 a n d 6 h
Maximal between 4 ancJ 8 h
,.,.,.,
,., ..,.
2nd nnt
ex
Commence from 24 h, extensive
Predominantly thrombotic; less inflamrnatc
Predominimtly inflamm less throrr.L-*:-
Hc?terophilsfor1med only a par t of the inflalmmatory re;action
lnflammatory reaction predominimtly neutrophilic from 1 to IB h
Monocyte!;(monocytic cellsf large mononuclea cells and niacrophages)
Participation of almost similar intensity right from the early stages
Little participation before 6 to 8 h
Lymhocytc (large and
Slight to moderate, predominantly perivascular lymphoid aggrergation from 3 to 8 h, and intense aggregation frcIm 12 h onwards
Only rarely in aggregates; more diffuIse and appe; at a much later stage (is from 12 h onwards)
Necrosis and icroscopic ch Vascular cha
-
m ,.
.mmnnrs 2 - . +
...
6 h, and
BaSOpnllS
gnificant pad:clpanon 2to4h
No significanr panlctpanon at 2 to 4 h
Eosinophil:
3t definite; d ifficult to sess
Significant in numbers at 24 to 48 h
lent of perivascular lymphoid aggregaIns; and 4, resemblance of AAR wit h delayedI persensitivity in the later stage!S. The most prominent feature of the re: iction was; '- -. e formation of thrombi in the early stages. a".~ n- i s is preceded by hyaline degeneration of the vessel ills. While the precise nature of the hyaline aterial remains unknown, it could be a conse.dence of antigen-antibody reaction (Jonsson 1972). In the chicken, thrombi were confined to the upper dermis away from areas of inflammation. It is possible that higher concentrations of immune re;actants in this region may attra.ct thrombocytes and in:itiate thrombosis. In the rabbit, thrombosis has be,en attributed to glycolytic prcoducts of leucocytes - - -. - - endothelium -. wllich are toxic to vascular (Stetson 19151), or to the damage effected by vascular inflam.. m;ation (Cochrane and Janoff 1974). The presenl: re:sults indicate that in the chicken thrombosis does; n o)t appear to be a consequence of inflammatory .scular dannage. The fibrin negative amorpkIOUS mate:rials seenI e in the lulmen of the: vessel are ling the wi311 and fre~ -..- - currlutelrrs 1 ,nL->..- .1.-~parently .immunt: turluuaorrar e-. l a1 ,cur in the 182). Similar deposit -
- -
-~~~
-1
-I
..,
reaction la^^^^. I firbar, l r a u r r a JU~;~;GJL that th rombus formation in the upper dermis begins with dc:position of immune complexes, adherence of throml3ocytes to the injured endothelium and their -. aggregation. It is also likely that the chicken thrombocyte may be engulfing immune complexes since it is established that it is a phagocytic cell (Glick et a1 1964, Carlson et a1 1968, Chang and Hamilton 1979, Awadhiya et a1 1980). The present stud:Y also revealed thrombocytes with engulfed carbon. Recently, in rabbit basophils, the presence of a platelet aggregating factor has been demons trated (Frick 1980). While nothing is known in the chicken, it may be significant that basophils were seen participating in the early Arthus reaction in considerable numbers and mav have a role in thrombocvte aggreg;ation. The inflammatory chanlges in th e chicken were confinced to the 1ower dernnis. -This nnight have: been . due to a low concentration of immune reactants in this region. As in the rabbit, the immune reactants may activate complement and produce factors chemotactic to leucocytes (Cochrane and Janoff N C,.U$
---
-
1974).
he chicke,n, the in~itial emig
comprised heterophils. This was soon followed by ing with various stimuli they reported the aggregates concurrent emigration of monocytic cells and occurring much later (Carlson and Allen 1970, basophils. A similar mixed exudate has been report ed Carlson 1972, Nair 1973, Awadhiya et al 1981a). by Chand et al (1976) in passive cut;aneous anla- Earl:y developnnent of lynnphoid aggregations in AAR phylaxis. Mixed cellular responses havse even been suggests it to ble a respon!;e to the antigenic stimulus. n _-.-._.: ~. ns Tt reported in non-immunoloeical inr~ammarlo --- -le lesions at ~4 and 48 hours histoloeicallv - u-----.-=----a (Carlson 1972, Nair 1973, A.wadhiya Iet a1 1981a, resernbled delayed hypersensitivity reactions (Ander1981b). The present results in1dicate thait the inflam- son 1971, Cheville and Richards 1971, War1ier et al matory changes in the chicker1 were lessI severe th an 1971, Rose and Bradley 1977). Despite the a pparent those in the rabbit. In tl.,h a latter neutrophils resemblance, AAR cannot be considered a!r3 oa +Li-..a dominated the early reaction (Gel1 and Hinde 195i4, dela!led hypersensitivity reaction iin the late1. stages, Humphrey 1955, Cochrane and Weigle 19513), asit is initiated by antigc:n-antiboc ly comple~ ces, and whereas in the sheep, there was delayed migration of its a:ssociation with cellurlar immu;nity has not been >.-. neutrophils and an early infiltration of mononuclear aernonstrated. cells (Vegad and Lancaster 1972, Vegad 1979). Bas on this work it appears that heterophils:,monocyl and basophils are involved in early inflixmmation 7 cllfiallthe chicken. Heterol:jhils were actively piiagocytic. In the fotlr- Werlke, Hanover, West Germany for the su~ P P ~ofY ~ O carbon. Our thanks are also due to I)r R. P. hour AAIR -in the by repeat ed C O ~ lidal . chicken, riensitised .Awa dhiya and the dean, Dr B. S. Malik f or their injections of antlgen in Freund's complete adjuva~ It, and to Dr A. G. Khan, senior scientis. almost all the heterophils in the exudate revealed Interest D,...l try Project, for the supply of b irds. phagocytosed carbon and the amorphous materi; Awadhiya et a1 (1980) demonstrated a lack of phag Recelived for pr ublication December cytic activitv . bv. hetero~hilsin earlv inflammatia Hence thle presenlt inding!r suggest that thleir phagocytic: propert ies may depend upon thleir References sensitisaticIn. The early appearance ot monocytes suggests their ANDERSON. J. C. (1971) Research in Veterinary Sc;nn- 1' 312-317 possible role in phagocytosis and degradation of R. P., VEGACI, J. L. & ULOLTE. G. immune complexes, thus supporting heterophils in AWADHIYA, Research in Veterenary Sciencc?29,120-122 the process. The monocytes appeared more uniform AWADHIYA, R. P.. VEGAD, I. L. & KiOLTE, G. b ".:.~.. Research in Vetennary acrence 31.23 1-235 in their phagocytic activity towards carbon and AWADHIYA, R. P.. VEGAD ULIE, cr. n. (IYUID) immune complexes, regardless of sensitisation. These Avian Pathology 10,3 13-320 findings are in contrast to those in the rabbit, in BENEDICT. A. A. (1%7) Methoas m immunology and Immunowhich the antigen is revealed by neutrophils in the chemistry, Vol 1. Eds C. A. Williams and M. W. Chase. New York, Academic Press, pp 229-234 early hours, but not by mononuclear cells (Cochrane et a1 1959). Further studies arc: obviousl! required to CARLSON, H. C. (1972)AvionDiseuses 16. ! H. C. & ALLEN, J. R. (1978 elucidate the functional signiificance o f monocytic CARLSON, 817-833 cells in avian inflammation. CARLSON, H. C., SWEENY, P. R. & TAKARYK, J. in Diseases 12,700-715 The basophils were not only seen intra- and extl 4D. N., CARLSON, H. (::. & EYRE, :P. (1976) Int~ vascularly, but were also found phagocytising carbl Arc hives of Allergy and Appli,ed Immunolorgy51,508-5117 particles. Dhodapkar et a1 (1982) described phagloCHAP4G, C. F . & HAMILT(IN, P. B. (:1979) Journa1 of the cytosis of carbon by basophils in reverse (passijre) P .~cuIoendothelial ~, -R.-. Society 25.-585-5'30 - -- - - Arthus reaction. This property of basoptlils has again CHEVILLE, N. F. & RICHARDS, W. D. (1971)Americom Journal of P a ~ h o l o g64,97-114 ~ been demonstrated. The present filndings thus COCHRANE. C. G. & JANOFF. A. (1974) The Infl endorse the possibility that tmsophils Inay share a Process, Vol 2. Eds B. W. Zweifach, L. Grant a1 phagocytic property with other cells under immuno.'"-'Jluskey. New York. Academic Press. py,OJ-I"L 68) Journal I logic stim~ 1lus. COCFIRANE. C. G. & WEIGL menrtolMedicine 108,591 404 Anothelr noteworthy feature of Ithe chicken DIXON. F. COCFIRANE. C. G., WEIGLI reaction 1was the formation of early perivasculIar -494 . . Jou rnol of Evperimenlal Medrctne nu, 481.. onnranot, P r n m 9 A hn.. rc n"..,"v .A* -... . lymphoid awjljlc5arca. I nvrrn UJ CULLING, C. F. A. (1974) Handbook of Histopatnological Techaggregates became more cornpact an(1 tended to niques. 3rd edn. London, Buttenvorths. p i N. (1982) coalesce. There was a prepon derance of monocytic DHODAPKAR, B. S., VEGAD, J. L. & 1 Research in VeterinaryScience 33,377-379 cells among cellular infiltrations. The ccurrence of FRICK, 0 . L. (1980) Basic and Clinical Immunology. EIds H. H. .> .-.~....--. 1s consistent - -- -2 ..: perivascular lymphoia aggregations wNn Fud enberg, D. P . Stites, J . L. Caldwell andI J. V. Wells. Maruzea. Lange Medical Publications. r,p 274-294 the findings of other investigaltors. Hovvever, WOI'k- 3
.,.,.
LA..
.I
* A .
! . A .
-2
e .
I",L\
or
-
AlWU L D
Wll"41
08
?.
I
L
L
-- - .
OC
lL3
,.
LOUGHBY. D. A. (1978:I InflammaticJn. Eds ;. H. Ferreura. New Yo1ik, Springer-Verlag.
I r
VEGAD VEGAD Veteri, WARNE notion
. . ..
k KANTOR,
) Inter-
nv 40.