Genetic Control of the Immune Response to Poly(GIu52Lys33Tyr15) in Neonatally Thymectomized High and Low Responder Rats Bridgett K. Davis, MD, John W. Shonnard, MD, and Thomas J. Gill 111, MD
The immune response to poly(Glu52Lys-Tvr"5) is under polygenic control and linked to the major histocompatibility complex of the rat. Aggregation of this antigen with methylated bovine serum albumin (MeBSA) eliminates the expression of genetic control by increasing the response of low responders and decreasing that of high responders. Humoral and cellular aspects of the immune response to both unaggregated and aggregated poly(Glu5"LvsnTvr"5) were investigated in neonatallv thvmectomized highresponder ACI and low-responder F344 rats. T cells are necessary for responses to unaggregated poly(Glu52Lys"Tyvr') since thymectomy significantly decreased numbers of antibodv-forming cells and serum antibodv levels, and delayed hypersensitivity responses and antigen-induced in vitro proliferation. However, thymectomy had no significant effect on these parameters of immune responsiveness in either ACI or F344 rats immunized with poly(Glu5"Lys3Tyr15)/.MeBSA. Aggregation also increased IgG production and delayed hypersensitivity and antibody affinity in low responders. (Am J Pathol 74:55-68, 1976)
COOPERATION OF BONE MARROW-DERIVED (B) and thvmus-derived (T) Iymphocytes is required for antibody responses to many antigens,' 2 but the production of IgNM immunoglobulin appears to be less dependent upon T cells than the production of IgG immunoglobulin.36 The nu/,nu mice, which lack thymuses, produce only IgNt.7 Studies of thymectomized mice and rats have demonstrated defects of humoral immunity, primarily characterized by a decrease in production of IgG immunoglobulin,6 Fllas wvell as a profound depression of expressions of cell-mediated immunity such as phytohemagglutinin (PHA) response and skin test reactivity.8"2 The immune response to both large particulate antigens, such as intact erythrocvtes, and natural and synthetic protein antigens depends on the presence of T cells. 1,2 A group of "T-independent" antigens characterized by repeating subunits within a large molecule, such as pneumococcal polvsaccharide, levan, polyvinylpyrrolidone and polymerized flagellin,'3-17 has been described. NMost T-independent From the Department of Pennsv hvania.
Pathology.
University of Pittsburgh School of Medicine. Pittsburgh.
Supported by Grants CA-18659 and GM-00135 from the National Institutes of Health: Dr Davis swas the recipient of Fellowship HD-00763 from the National Institutes of Health. and Dr Shonnard swas a Sarah Scaife Fellow- in Pathology. Accepted for publication March 13. 1976 Address reprint requests to Dr. Thomas J Gill III. Department of Pathology. Scaife Hall. U niversity of Pittsburgh. Pittsburgh. PA 13213. 55
56
DAVIS ET AL
American Journal of Pathology
antigens stimulate IgM antibodies,18 although occasional T-independent antigens do induce an IgG response.1' The immune response to many natural and svnthetic antigens is under genetic control and is linked to the major histocompatibility complex (M1HC) of several species."` Genetically nonresponding mice appear to be functionally thvmectomized, since they produce only IgM antibody to a synthetic polypeptide to which high responders form both IgM and IgG.' Thymectomized high responder mice also produce onlv IgM.6 Extensive investigation utilizing inbred rats has characterized the polygenic control of the immune response to the synthetic polypeptide polv(Glu52Lvs3"Tvr5)24 and its linkage to the major histocompatibilitv locus.25 " ACI rats are representative of high responders, while F344 rats are representative of low responders. Aggregation of poly(Glu52Lvs&¶TvrlS) with methvlated bovine serum albumin (MeBSA) increases the response in low responders while decreasing that of high responders.2" ACI rats, when immunized with either aggregated or unaggregated antigen, are good producers of both IgG and IgM immunoglobulins. F344 rats, on the other hand, produce little antibodv to unaggregated polv(Glu52LyLvsTyrl5), and thev make IgG with verv little IgM-antibody to
poly(Glu52 Lys3Tvtr15 )/ NMeBSA.
Since IgG production is influenced by T cells to a greater extent than IgM production, and since one significant difference between the immune response to polv(Glu52Lvs"Tvr`5) of high and low responder rats is expressed at the level of IgM and IgG synthesis, studies utilizing neonatallv thymectomized rats were performed to a) document the need for T cells in the antibody response to polI(Glu52LLs"Tyr15), b) investigate T-cell function in high and low responder strains, and c) evaluate the effects of aggregation of poly(Glu52Lvs3sTyrv5) with MeBSA on the immune response of animals with decreased T-cell populations.
Materials and Mefthos An*nals Inbred ACI and F344 rats mvere obtained either from Microbiological Associates, Walkersv ille. Md., or from our own breeding colony. They w ere maintained on Purina rat chow and mvater ad libitumn and were routinely mated at 8 to 10 weeks of age.
Thmctomies and Sham-Thar*ctomies Neonatal rats mvere th\mectomized or sham-thvmectomized within :36 hours after birth according to the method of Jankovic et al.8 Briefly. the young rats were anesthetized in an ice wlater bath until respiration and movement had ceased. A median stemal-splitting incision %vas made, and the thymus mvas gently eased out of the thorax. Silk sutures were used to approximate the ribs and skin. Sham-thvmectomies were treated in the same was;
Vol. 84, No. 1
July 1976
GENETIC CONTROL OF IMMUNE RESPONSE
57
the th% mus w-as exposed but left intact. The animals were placed in a closed container until warm and then retumed to the mother. Th%-mectomized female offspring were utilized in all studies, and thesy were immunized w-hen 8 to 12 weeks old.
Anlti and ImmunitI The antigen w-as poly(Glu52Lvs-Tv r`). X -hich is a random linear sy nthetic poly peptide with a molecular weight of 28,000 daltons. Mlethvlated bosine serum albumin was prepared according to Sueoka and Cheng.9 Aggregated antigen was made by adding an equivalent weight of MeBSA in water to antigen in water. as previouslv described.-' All animals received an immunizing dose of 0. 73 mg polv(Glu52LyssTyr'5) or polv (Glu52L!s`Tyr15) MeBSA (0.73 mg antigen and 0.75 mg MeBSA) in a total of 0.3 ml of Freund's complete adjuvant following the standard protocol (0.13 ml in each hind footpad, 0.2 ml in the subcutaneous tissue at the back of the neck).' The second injection of 0.73 mg of the appropriate antigen was given in w-ater in the peritoneal cavitx 6 days later for hemolx-tic plaque-forming cell (PFC) assays and 21 days later for in vitro antigen stimulation studies. Nineteen davs after the first immunization. groups of animals received intradermal injections of 100 gg of poly (Glu52Ly s`"Ty r'5). poly(Glu52LLys'3Ty r5 MeBSA. or MeBSA alone in 0.05 ml of 0.13 N1 NaCl. Fortv-eight hours later, the test sites were graded by diameter of induration (l+. I to 3 mm. 2+. 4 to 6 mm; :3+. 7 to 7 mm: 4+. 10 to 12 mm) and excised for histologic examination. Similar skin tests were always negative in unimmunized animals.
Anibody and Antbody-Fming Cells Serum antibody \vas quantitated by the bromoacetyl cellulose immunoadsorbent micromethod "\w-hich can detect 10 jig antibodv ml. For PFC assavs. all animals receised a second injection of 0. 735 mg of the appropriate antigen 6 days follom.-ing the initial immunization. and the l!-mphoid organs w-ere harsested 4 davs thereafter; this time w-as previously determined to be the peak of the response.' PFC from purified popliteal lymph node cells were determined utilizing the Cunningham liquid monolay er modification of the Jeme hemoly tic plaque technique with a mixture containing equal amounts of sheep red blood cells (SRBC) coupled wvith pol\ (Glu52LLsUTyr'5). rat ly mph node cells, guinea pig serum as a source of complement. and for indirect (IgG) plaque dev elopment. rabbit antirat IgG at a final dilution of 1: 256.-" The rabbit antirat antisera %vas absorbed three times with SRBC to remove agglutinin activitv and \-as specific by immunodiffusion. No detectable light chain activity w-as present. The variability in the PFC assay sy stem among the animals s\ithin each group was great. For this reason, differences of P < 0.001 \vere considered highly significant and P < 0.0-2. of border line significance. A value of 30 PFC 10 w-as presiouslv determined to be background level.' In Vitro StWies
Isolated spleen cells were cultured in Medium-199 with Hepes buffer (40 mnM). 3 X 10-5 Mg ml)-penicillin (100 units ml) supplemented X ith 10%c heat-inactivated ACI rat serum in a total volume of 1.0 ml.'4 Cultures were performed in triplicate in 12 X 75 mm poly propylene tubes (Falcon Plastics, Los Angeles. Calif.). Phytohemagglutinin (PHA-P) concentrations ranged from 0 to :3.2 Ml PHA-P ml. and these cultures, containing 1.3 x 108 cells, were terminated on the third dav. Cultures for antigen stimulation contained 3.0 X 108 cells and 100 gg of polv (Glu52yL s"Ty r'5). \eBSA. or polv (Glu52L ys-UTy r') aggregated X ith \MeBSA. and the cultures mvere harvested on the fourth day. Approximately 18 hours prior to termination. all cultures were pulsed with 2 MCi 3H-methvl thvmidine. harvested with a Skatron multihan-ester (Flow Laboratories. Rockville. \Id.). dried and counted in liquid scintillation \1 2-mercaptoethanol. and streptomrncin (100
counter.
58
DAVIS ET AL
American Journal of Pathology
Lymphocyte Surface Markers Lymphocytes from spleens and lmph nodes of thvmectomized and sham-thNmectomized ACI and F344 rats w-ere isolated on Ficoll-Hypaque gradients (one part 40%S Hy paque to two parts 9% Ficoll) in order to determine the percentage of B and T cells in each population. The cells w-ere swashed three times in \1-199. incubated wsith rabbit antirat IgG or rabbit anti-rat brain antisera, and washed again. They wsere then incubated w-ith goat antirabbit IgG conjugated with fluorescein. washed again, and examined in 45% gly cerol. The antisera were prepared in rabbits by a series of intramuscular injections of purified rat IgG or F344 rat brain.5" Anti-IgG Nvas absorbed w-ith SRBC to remove agglutinin activitv and ssas specific for IgG by immunodiffusion swith no evidence of light chain activitv. Antibrain antisera absorbed with F344 RBCs was cytotoxic to a dilution of 1: 160 and stained 99% of thymocytes from both ACI and F344 rats.
Results Antbody Formation
Thvmectomy markedly decreased both direct (1g1) and indirect (IgG) PFC in ACI rats immunized with polv(Glu52Lvs'"Tvr`). Sham-thvmectomized F344 rats produced extremely low levels of direct and indirect PFC to unaggregated polv(Glu52Lys33T-r`5), and thymectomv did not alter the response in the F344 rats (Table 1). However, when polv(GClu52Lvs3"Tvr'5) aggregated with MeBSA was the immunizing antigen, thymectomv had no significant effect on the indirect (IgG) PFC response and only a questionable effect on the direct (IgNM) PFC response (P < 0.02) in either the ACI or F344 strain (Table 1). Serum antibody levels also reflected the effect of thymectomy. The only significant difference occurred in thmectomized ACI animals, w-hich, followving immunization with pol-(Glu52Ly-s3Tyr'5), had decreased serum antibody levels compared to sham-thy-mectomized ACI rats (P < 0.001) (Table 2). Cellular Immunrt
Delayed hypersensitivity was evaluated after immunization with either polv(Glu52Lvs-"Tyr'5) or polv(Glu52Lys33Txr'5) aggregated with MeBSA followed by intradermal testing with the appropriate antigen 19 days later (Table 3). Sham-thvmectomized ACI rats immunized with poly(Glu52Lys3Tyr'5) had good skin test responses, wvith most reacting in the 4+ range, thy-mectomy- abolished or markedly reduced this response. FP344 rats, both sham-thv-mectomized and thv-mectomized, had no delav ed hypersensitivity response followving immunization wvith poly-(Glu52vLys33Tvr'5). Thymectomv did not alter significantly the pattern of delayed hypersensitivity in either the ACI or F344 strains immunized ssith polv(Glu52Lyss33Tvr'5) fMeBSA. ACI rats immunized wvith polv-(Glu52Lvs3T-r'5) MeBSA had good delayed hypersensitivity re-
Vol. 84, No. 1 July 1976
GENETIC CONTROL OF IMMUNE RESPONSE
59
Table 1-Plaque-Forming Cells per 10' Popliteal Lymph Node Cells in Thymectomized and Sham-Thymectomized Female ACI and F344 Rats Strain
Procedure
Direct PFC (mean ± SE)
Indirect PFC (mean ± SE)
17 ± 5 220 ±67 (P
-n
~CII
4
CM
CO
5
_
co
0O
CV o r~ ul) co r_1- v_ _
X
C)
E
+-+-+ +-H -H++ 0) 0 Oco 0I) CDC0(0 c C) CqwJ0)
0
OL
CM o0C) owt co CNI lt CV CV U) LO
c
-
cn
V) CI) cm
C
c.1 0 C
0 CD
c
)JC
C M
CD
CD
0
0)
>OE CD >
_0 --
>%
C
0
E
0-
m
CNl
E 0 0
0 0
m 0 0
0
0 >,0
-
0
0
0E
L-
In
03
o_
x 0O 0
.
0E 'aC
co o
0
r-
cc0
a. CID
'E< < E
0
0
0 0 0
0 cm
E.
-5.
cE-
E E
The immune response to poly(Glu52Lys-Tvr"5) is under polygenic control and linked to the major histocompatibility complex of the rat. Aggregation of this antigen with methylated bovine serum albumin (MeBSA) eliminates the expression of genetic control by increasing the response of low responders and decreasing that of high responders. Humoral and cellular aspects of the immune response to both unaggregated and aggregated poly(Glu5"LvsnTvr"5) were investigated in neonatallv thvmectomized highresponder ACI and low-responder F344 rats. T cells are necessary for responses to unaggregated poly(Glu52Lys"Tyvr') since thymectomy significantly decreased numbers of antibodv-forming cells and serum antibodv levels, and delayed hypersensitivity responses and antigen-induced in vitro proliferation. However, thymectomy had no significant effect on these parameters of immune responsiveness in either ACI or F344 rats immunized with poly(Glu5"Lys3Tyr15)/.MeBSA. Aggregation also increased IgG production and delayed hypersensitivity and antibody affinity in low responders. (Am J Pathol 74:55-68, 1976)
COOPERATION OF BONE MARROW-DERIVED (B) and thvmus-derived (T) Iymphocytes is required for antibody responses to many antigens,' 2 but the production of IgNM immunoglobulin appears to be less dependent upon T cells than the production of IgG immunoglobulin.36 The nu/,nu mice, which lack thymuses, produce only IgNt.7 Studies of thymectomized mice and rats have demonstrated defects of humoral immunity, primarily characterized by a decrease in production of IgG immunoglobulin,6 Fllas wvell as a profound depression of expressions of cell-mediated immunity such as phytohemagglutinin (PHA) response and skin test reactivity.8"2 The immune response to both large particulate antigens, such as intact erythrocvtes, and natural and synthetic protein antigens depends on the presence of T cells. 1,2 A group of "T-independent" antigens characterized by repeating subunits within a large molecule, such as pneumococcal polvsaccharide, levan, polyvinylpyrrolidone and polymerized flagellin,'3-17 has been described. NMost T-independent From the Department of Pennsv hvania.
Pathology.
University of Pittsburgh School of Medicine. Pittsburgh.
Supported by Grants CA-18659 and GM-00135 from the National Institutes of Health: Dr Davis swas the recipient of Fellowship HD-00763 from the National Institutes of Health. and Dr Shonnard swas a Sarah Scaife Fellow- in Pathology. Accepted for publication March 13. 1976 Address reprint requests to Dr. Thomas J Gill III. Department of Pathology. Scaife Hall. U niversity of Pittsburgh. Pittsburgh. PA 13213. 55
56
DAVIS ET AL
American Journal of Pathology
antigens stimulate IgM antibodies,18 although occasional T-independent antigens do induce an IgG response.1' The immune response to many natural and svnthetic antigens is under genetic control and is linked to the major histocompatibility complex (M1HC) of several species."` Genetically nonresponding mice appear to be functionally thvmectomized, since they produce only IgM antibody to a synthetic polypeptide to which high responders form both IgM and IgG.' Thymectomized high responder mice also produce onlv IgM.6 Extensive investigation utilizing inbred rats has characterized the polygenic control of the immune response to the synthetic polypeptide polv(Glu52Lvs3"Tvr5)24 and its linkage to the major histocompatibilitv locus.25 " ACI rats are representative of high responders, while F344 rats are representative of low responders. Aggregation of poly(Glu52Lvs&¶TvrlS) with methvlated bovine serum albumin (MeBSA) increases the response in low responders while decreasing that of high responders.2" ACI rats, when immunized with either aggregated or unaggregated antigen, are good producers of both IgG and IgM immunoglobulins. F344 rats, on the other hand, produce little antibodv to unaggregated polv(Glu52LyLvsTyrl5), and thev make IgG with verv little IgM-antibody to
poly(Glu52 Lys3Tvtr15 )/ NMeBSA.
Since IgG production is influenced by T cells to a greater extent than IgM production, and since one significant difference between the immune response to polv(Glu52Lvs"Tvr`5) of high and low responder rats is expressed at the level of IgM and IgG synthesis, studies utilizing neonatallv thymectomized rats were performed to a) document the need for T cells in the antibody response to polI(Glu52LLs"Tyr15), b) investigate T-cell function in high and low responder strains, and c) evaluate the effects of aggregation of poly(Glu52Lvs3sTyrv5) with MeBSA on the immune response of animals with decreased T-cell populations.
Materials and Mefthos An*nals Inbred ACI and F344 rats mvere obtained either from Microbiological Associates, Walkersv ille. Md., or from our own breeding colony. They w ere maintained on Purina rat chow and mvater ad libitumn and were routinely mated at 8 to 10 weeks of age.
Thmctomies and Sham-Thar*ctomies Neonatal rats mvere th\mectomized or sham-thvmectomized within :36 hours after birth according to the method of Jankovic et al.8 Briefly. the young rats were anesthetized in an ice wlater bath until respiration and movement had ceased. A median stemal-splitting incision %vas made, and the thymus mvas gently eased out of the thorax. Silk sutures were used to approximate the ribs and skin. Sham-thvmectomies were treated in the same was;
Vol. 84, No. 1
July 1976
GENETIC CONTROL OF IMMUNE RESPONSE
57
the th% mus w-as exposed but left intact. The animals were placed in a closed container until warm and then retumed to the mother. Th%-mectomized female offspring were utilized in all studies, and thesy were immunized w-hen 8 to 12 weeks old.
Anlti and ImmunitI The antigen w-as poly(Glu52Lvs-Tv r`). X -hich is a random linear sy nthetic poly peptide with a molecular weight of 28,000 daltons. Mlethvlated bosine serum albumin was prepared according to Sueoka and Cheng.9 Aggregated antigen was made by adding an equivalent weight of MeBSA in water to antigen in water. as previouslv described.-' All animals received an immunizing dose of 0. 73 mg polv(Glu52LyssTyr'5) or polv (Glu52L!s`Tyr15) MeBSA (0.73 mg antigen and 0.75 mg MeBSA) in a total of 0.3 ml of Freund's complete adjuvant following the standard protocol (0.13 ml in each hind footpad, 0.2 ml in the subcutaneous tissue at the back of the neck).' The second injection of 0.73 mg of the appropriate antigen was given in w-ater in the peritoneal cavitx 6 days later for hemolx-tic plaque-forming cell (PFC) assays and 21 days later for in vitro antigen stimulation studies. Nineteen davs after the first immunization. groups of animals received intradermal injections of 100 gg of poly (Glu52Ly s`"Ty r'5). poly(Glu52LLys'3Ty r5 MeBSA. or MeBSA alone in 0.05 ml of 0.13 N1 NaCl. Fortv-eight hours later, the test sites were graded by diameter of induration (l+. I to 3 mm. 2+. 4 to 6 mm; :3+. 7 to 7 mm: 4+. 10 to 12 mm) and excised for histologic examination. Similar skin tests were always negative in unimmunized animals.
Anibody and Antbody-Fming Cells Serum antibody \vas quantitated by the bromoacetyl cellulose immunoadsorbent micromethod "\w-hich can detect 10 jig antibodv ml. For PFC assavs. all animals receised a second injection of 0. 735 mg of the appropriate antigen 6 days follom.-ing the initial immunization. and the l!-mphoid organs w-ere harsested 4 davs thereafter; this time w-as previously determined to be the peak of the response.' PFC from purified popliteal lymph node cells were determined utilizing the Cunningham liquid monolay er modification of the Jeme hemoly tic plaque technique with a mixture containing equal amounts of sheep red blood cells (SRBC) coupled wvith pol\ (Glu52LLsUTyr'5). rat ly mph node cells, guinea pig serum as a source of complement. and for indirect (IgG) plaque dev elopment. rabbit antirat IgG at a final dilution of 1: 256.-" The rabbit antirat antisera %vas absorbed three times with SRBC to remove agglutinin activitv and \-as specific by immunodiffusion. No detectable light chain activity w-as present. The variability in the PFC assay sy stem among the animals s\ithin each group was great. For this reason, differences of P < 0.001 \vere considered highly significant and P < 0.0-2. of border line significance. A value of 30 PFC 10 w-as presiouslv determined to be background level.' In Vitro StWies
Isolated spleen cells were cultured in Medium-199 with Hepes buffer (40 mnM). 3 X 10-5 Mg ml)-penicillin (100 units ml) supplemented X ith 10%c heat-inactivated ACI rat serum in a total volume of 1.0 ml.'4 Cultures were performed in triplicate in 12 X 75 mm poly propylene tubes (Falcon Plastics, Los Angeles. Calif.). Phytohemagglutinin (PHA-P) concentrations ranged from 0 to :3.2 Ml PHA-P ml. and these cultures, containing 1.3 x 108 cells, were terminated on the third dav. Cultures for antigen stimulation contained 3.0 X 108 cells and 100 gg of polv (Glu52yL s"Ty r'5). \eBSA. or polv (Glu52L ys-UTy r') aggregated X ith \MeBSA. and the cultures mvere harvested on the fourth day. Approximately 18 hours prior to termination. all cultures were pulsed with 2 MCi 3H-methvl thvmidine. harvested with a Skatron multihan-ester (Flow Laboratories. Rockville. \Id.). dried and counted in liquid scintillation \1 2-mercaptoethanol. and streptomrncin (100
counter.
58
DAVIS ET AL
American Journal of Pathology
Lymphocyte Surface Markers Lymphocytes from spleens and lmph nodes of thvmectomized and sham-thNmectomized ACI and F344 rats w-ere isolated on Ficoll-Hypaque gradients (one part 40%S Hy paque to two parts 9% Ficoll) in order to determine the percentage of B and T cells in each population. The cells w-ere swashed three times in \1-199. incubated wsith rabbit antirat IgG or rabbit anti-rat brain antisera, and washed again. They wsere then incubated w-ith goat antirabbit IgG conjugated with fluorescein. washed again, and examined in 45% gly cerol. The antisera were prepared in rabbits by a series of intramuscular injections of purified rat IgG or F344 rat brain.5" Anti-IgG Nvas absorbed w-ith SRBC to remove agglutinin activitv and ssas specific for IgG by immunodiffusion swith no evidence of light chain activitv. Antibrain antisera absorbed with F344 RBCs was cytotoxic to a dilution of 1: 160 and stained 99% of thymocytes from both ACI and F344 rats.
Results Antbody Formation
Thvmectomy markedly decreased both direct (1g1) and indirect (IgG) PFC in ACI rats immunized with polv(Glu52Lvs'"Tvr`). Sham-thvmectomized F344 rats produced extremely low levels of direct and indirect PFC to unaggregated polv(Glu52Lys33T-r`5), and thymectomv did not alter the response in the F344 rats (Table 1). However, when polv(GClu52Lvs3"Tvr'5) aggregated with MeBSA was the immunizing antigen, thymectomv had no significant effect on the indirect (IgG) PFC response and only a questionable effect on the direct (IgNM) PFC response (P < 0.02) in either the ACI or F344 strain (Table 1). Serum antibody levels also reflected the effect of thymectomy. The only significant difference occurred in thmectomized ACI animals, w-hich, followving immunization with pol-(Glu52Ly-s3Tyr'5), had decreased serum antibody levels compared to sham-thy-mectomized ACI rats (P < 0.001) (Table 2). Cellular Immunrt
Delayed hypersensitivity was evaluated after immunization with either polv(Glu52Lvs-"Tyr'5) or polv(Glu52Lys33Txr'5) aggregated with MeBSA followed by intradermal testing with the appropriate antigen 19 days later (Table 3). Sham-thvmectomized ACI rats immunized with poly(Glu52Lys3Tyr'5) had good skin test responses, wvith most reacting in the 4+ range, thy-mectomy- abolished or markedly reduced this response. FP344 rats, both sham-thv-mectomized and thv-mectomized, had no delav ed hypersensitivity response followving immunization wvith poly-(Glu52vLys33Tvr'5). Thymectomv did not alter significantly the pattern of delayed hypersensitivity in either the ACI or F344 strains immunized ssith polv(Glu52Lyss33Tvr'5) fMeBSA. ACI rats immunized wvith polv-(Glu52Lvs3T-r'5) MeBSA had good delayed hypersensitivity re-
Vol. 84, No. 1 July 1976
GENETIC CONTROL OF IMMUNE RESPONSE
59
Table 1-Plaque-Forming Cells per 10' Popliteal Lymph Node Cells in Thymectomized and Sham-Thymectomized Female ACI and F344 Rats Strain
Procedure
Direct PFC (mean ± SE)
Indirect PFC (mean ± SE)
17 ± 5 220 ±67 (P
-n
~CII
4
CM
CO
5
_
co
0O
CV o r~ ul) co r_1- v_ _
X
C)
E
+-+-+ +-H -H++ 0) 0 Oco 0I) CDC0(0 c C) CqwJ0)
0
OL
CM o0C) owt co CNI lt CV CV U) LO
c
-
cn
V) CI) cm
C
c.1 0 C
0 CD
c
)JC
C M
CD
CD
0
0)
>OE CD >
_0 --
>%
C
0
E
0-
m
CNl
E 0 0
0 0
m 0 0
0
0 >,0
-
0
0
0E
L-
In
03
o_
x 0O 0
.
0E 'aC
co o
0
r-
cc0
a. CID
'E< < E
0
0
0 0 0
0 cm
E.
-5.
cE-
E E
