bluuebunny/crossref_metadata_2025 huggingface dataset:
STUDIES ON THE SENSITIZATION OF ANIMALS WITH SIMPLE CHEMICAL COMPOUNDS
Authors: Macher Egon , Chase Merrill W.
Published: June 2004

The fate of 14C-labeled allergens injected intradermally into guinea pigs, namely picryl chloride (PCl*) and 2:4 dinitrochlorobenzene (DNCB*), was followed during the induction period of delayed hypersensitivity. Both chemicals were applied in a single injection into one ear in amounts that approached their minimal sensitizing doses (PCl, 0.25 µg; DNCB, 5.0 µg). Radioactivity in the various tissues was determined by liquid scintillation counting after combustion of tissues to CO2 and H2O.

The injected allergens seemed to leave the injection site in three phases. A large proportion of allergen escaped rapidly from the ear, about 50% within 3 hr in the case of PCl, within 15 min for DNCB, the difference probably reflecting their unequal reaction constants. Initially there was a "half-life" escape in 2.5 hr with injected dosage of 0.25 µg PCl and in 18 hr for 5.0 µg DNCB. This escape occurred via the regional veins and not via the lymphatics. Radioactive decomposition products of the allergens were already present in the urine within 3–4 hr.

After 6–8 hr, the half-life time of escape lengthened to approximately 28 hr for both allergens used in their respective initial dosages, holding up to 2 days after which there occurred still further slowing; between 2 and 4 days the time was about 43 hr for PCl, much longer (72–88 hr?) for DNCB, apparently reflecting different physicochemical properties of this second fraction. Sensitization seemed to be connected with the portion that was present between 12 hr and 4 days of the induction period. It is not known how far the escape of radioactivity during this period may represent gradual hydrolysis of attached picryl and dinitrophenyl groupings, respectively, to form picric acid and dinitrophenol. Gradual accumulation of the second fraction in the regional lymph nodes could definitely be excluded. It was noted that no hypersensitivity arose and essentially no depot of radioactivity existed between 12 hr and 4 days when DNCB was injected in a dose of 0.25 µg, owing to its ready escape from the ear; but 20 times as much DNCB caused sensitization and provided about the same fixed depot as 0.25 µg of picryl chloride.

After delayed hypersensitivity had been established, traces of radioactivity were still measurable at the site. This third fraction, probably representing a different coupling product, escaped at a very low rate and was traceable up through several weeks.

No demonstrable radioactivity could be detected in thymus, spleen, and mesenteric nodes when examined at short intervals between ½ min and 17 days.

In analogy with findings on transplantation "immunity" and with studies reported in the following paper, the induction of delayed hypersensitivity can be explained by encounters between lymphoid cells and the hapten complex which is found present in the local site for 4 days, in agreement with Medawar's concept of peripheral sensitization.
THE HUMAN SECRETORY IMMUNOGLOBULIN SYSTEM: IMMUNOHISTOLOGICAL LOCALIZATION OF γA, SECRETORY "PIECE," AND LACTOFERRIN IN NORMAL HUMAN TISSUES
Authors: Tourville D. R. , Adler R. H. , Bienenstock J. , Tomasi T. B.
Published: June 2004

The immunohistological localization of γA, secretory "piece" (SP), and lactoferrin (LF) in the mucosae of a variety of normal human tissues was investigated using specific fluoresceinated antisera. γA staining was localized in the apical portion of the mucosal epithelium, intercellular spaces, basement membrane area, and plasma cells of the interstitium or lamina propria of a number of normal human tissues.

SP was ubiquitous in the mucosal epithelium of all tissues studied which included parotid and submaxillary glands, bronchi, pancreas, GI tract, sweat glands, kidney, and gall bladder. In addition, SP staining was localized in the intercellular spaces and on the surface of the epithelial cells lining the lumen of the secretory glands. No SP staining was observed in the plasma cells of the interstitium or lamina propria surrounding the secretory glands in these tissues, and no SP staining was observed in sections of normal spleen or lymph node tissue. SP staining was observed in the sweat glands, pancreas, and kidney in the absence of γA staining.

LF was much less ubiquitous in the epithelial cells of the various tissues studied and appeared to be restricted primarily to the acinar epithelium of the bronchial mucosae, parotid, and submaxillary salivary glands, and was also found in renal tubular cells.

A hypothetical model for the transport of γA and SP across mucosal membrane epithelium is presented.
REGULATION OF ANTIBODY SYNTHESIS AGAINST ESCHERICHIA COLI ENDOTOXIN
Authors: Britton Sven
Published: June 2004

Normal mouse lymphoid cells have been shown to become specifically paralyzed after in vitro exposure to high doses of detoxified endotoxin of Escherichia coli 055:B5. The immune status of the treated cells was tested after transfer to secondary irradiated hosts. Paralysis was shown to be initiated by events taking place in vitro, since the amount of antigen retained on the cells after the in vitro exposure was insufficient to induce paralysis in vivo. The induction of paralysis was dependent on the concentration of antigen added to the cells in vitro. Certain variables, such as time of exposure and temperature at exposure, influenced the ease by which the cells could be paralyzed. Cells pretreated with trypsin were not susceptible to induction of paralysis by the above procedure.
ESTERASES OF THE POLYMORPHONUCLEAR LEUKOCYTE CAPABLE OF HYDROLYZING ACETYL DL-PHENYL-ALANINE ß-NAPHTHYL ESTER
Authors: Becker Elmer L. , Ward Peter A.
Published: June 2004

Previous published work has led to the hypothesis that the activatable esterase of chemotaxis is a serine esterase of the rabbit polymorphonuclear leukocyte existing in an inert, phosphonate insusceptible form, which after activation is capable of hydrolyzing aromatic amino acid esters and being inhibited by phosphonates. In the present study, directed to the testing of this hypothesis, we have shown that rabbit peritoneal polymorphonuclear leukocytes contain three esterases capable of hydrolyzing the aromatic amino acid ester, acetyl DL-phenylalanine ß-naphthyl ester. Two of these esterases, esterase 1 and esterase 2, are inhibited by various p-nitrophenyl ethyl phosphonate esters. The inhibition of each esterase is irreversible and progressive with time. When the logarithm of the esterase activity remaining after cell and inhibitor have been in contact for a constant time is plotted against the concentration of inhibitor, a straight line results. These results support the conclusion that both esterases are serine esterases. The third esterase, esterase 3, differs from the other two by its inability to be inactivated by any of the phosphonates no matter how high the concentration of phosphonate or prolonged the period of incubation of cell with phosphonate.

The activity of esterase 1 is at least 10,000 times more easily inhibited by phosphonates than is that of esterase 2; incubating rabbit polymorphonuclear leukocytes for 15 min at 27°C with 10–9–10–8 M concentrations of various phosphonates inactivates esterase 1, but it required 10–6–10–4 M concentrations of the same phosphonates to inhibit esterase 2. The inhibition profiles of esterase 1 are distinctly different from those of esterase 2 when the two esterases are tested with the phenylalkylphosphonates, chloroalkylphosphonates, and alkylphosphonates.

The inhibition profile of esterase 1 is essentially the same as that of the activatable esterase of chemotaxis obtained previously when the same three homologous series of phosphonates were tested for their ability to protect against deactivation by the chemotactic factor or give chemotactic-dependent inhibition.

It is tentatively concluded that esterase 1 of the rabbit peritoneal neutrophil is the activated form of the activatable esterase of chemotaxis.
STUDIES ON THE IMMUNOTHERAPY OF RUNT DISEASE IN RATS
Authors: Silvers Willys K. , Billingham R. E.
Published: June 2004

Using rats of the Lewis and BN (Ag-B locus incompatible) isogenic strains, a comparative study has been made of the capacity to prevent or mitigate the development of runt disease with: (a) lymph node cell suspensions from normal adult BN rats, (b) node cells, or (c) serum from donors sensitized against Lewis tissue antigens, or (d) heterologous anti-lymphocyte serum (ALS) raised in rabbits against rat thymocytes.

Following a standard intravenous or intraperitoneal inoculation of 20 x 106 Lewis node cells into neonatal BN hosts, there are cutaneous manifestations of runt disease within 11–15 days and death invariably takes place within 20 days. However, complete protection is afforded by administration of a similar number of normal BN node cells via a different vein, or admixed with the otherwise harmful Lewis node cells. However, timing of the administration was crucially important—precedence or delay by as little as 4 hr resulted in a great impairment of protection. When the inoculations of the two cell suspensions were separated by 24 hr, no protection was afforded. These and other observations suggested that a necessary condition for protection of the hosts by unsensitized isologous cells requires that they establish a prompt and intimate confrontation with the homologous target cells.

At the same dosage level, suspensions of node cells from sensitized isologous donors were much more effective therapeutically, saving the lives of 92% of treated subjects when administered after a delay of 3 days, and of 19% when the delay was 4 or 5 days.

Of the various immunotherapeutic agents studied, daily injections of 0.2 ml of isoantiserum gave the best results, and could totally reverse the course of the disease even when initiated at age 10–13 days and subjects already presented symptoms. ALS, although inferior to isoantiserum at the dosage levels tested, proved to be superior to sensitized isologous cells as a protective agent, since the initiation of daily injections after delays of 6 or 8 days were still effective.

The observations that delayed treatments of infant rats with isoantisera or ALS resulted in complete recoveries sustain the thesis that the lesions responsible for the fatal outcome of runt diseases are not inflicted at a very early stage. The efficacy of both isoantisera and ALS as a means of inhibiting the progression of homologous disease also suggests that they may have therapeutic value in situations where this condition is encountered.
RELATION BETWEEN EPSTEIN-BARR VIRAL AND CELL MEMBRANE IMMUNOFLUORESCENCE IN BURKITT TUMOR CELLS
Authors: Pearson G. , Klein G. , Henle G. , Henle W. , Clifford P.
Published: June 2004

Previous reports (1, 2) have established that the expression of certain distinctive membrane antigen(s) on the surface of Burkitt's lymphoma (BL) and infectious mononucleosis (IM) cells is dependent on the presence of Epstein-Barr virus (EBV) in the cell line. The investigations reported here provide evidence that antibodies directed against EBV antigens, as revealed by the immunofluorescence test on acetone-fixed smears (8), and the membrane reactive antibodies, although often present in the same serum, are nevertheless distinctly different. Absorption of Mutua serum, the standard reference serum for demonstrating membrane antigen(s) on BL and IM cells, with BL cells completely removed anti-membrane activity without significantly affecting the anti-EBV antibody titer. Furthermore, sera were found which contained one type of antibody but not the other. Sera with high anti-membrane but low anti-EBV activity were found among relatives of BL patients. These sera reacted with the membranes of EBV-carrying BL and IM cells in essentially the same way, i.e., against the same spectrum of target cells, as the EBV-positive Mutua serum. They were unable to block the membrane reactivity of FITC-conjugated Mutua serum, however. In some cases they showed weak but incomplete blocking. One such EBV-negative, membrane-positive BL relative serum (Robert) was conjugated with FITC and used for direct staining of BL and IM cells. Again, this conjugate reacted against the same target cell spectrum as a Mutua conjugate, but its reactivity was completely blocked by a number of Burkitt patients' sera, although unconjugated Robert serum did not block the Mutua-conjugate. A number of other membrane-positive BL relative sera also failed to block Mutua, but completely blocked the Robert conjugate. A number of Swedish and African control sera and an isoantiserum gave no blocking against Robert conjugate. It therefore appears that the Mutua conjugate contains at least two antibody specificities against the EBV-determined membrane antigens. One, but not the other, is shared with the antibody specificity present in Robert's serum and a number of other sera from relatives of BL patients.
CELLS INVOLVED IN THE IMMUNE RESPONSE
Authors: Abdou Nabih I. , Richter Maxwell
Published: June 2004

Irradiated rabbits given allogeneic bone marrow cells from normal adult donors responded to an injection of sheep red blood cells by forming circulating antibodies. Their spleen cells were also capable of forming many plaques using the hemolysis in gel technique, and were also capable of undergoing blastogenesis and mitosis and of incorporating tritiated thymidine upon exposure to the specific antigen in vitro. However, irradiated rabbits injected with allogeneic bone marrow obtained from rabbits injected with sheep red blood cells 24 hr prior to sacrifice (primed donors) were incapable of mounting an immune response after stimulation with sheep red cells. This loss of reactivity by the bone marrow from primed donors is specific for the antigen injected, since the immune response of the irradiated recipients to a non-cross-reacting antigen, the horse red blood cell, is unimpaired. Treatment of the bone marrow donors with high-titered specific antiserum to sheep red cells for 24 hr prior to sacrifice did not result in any diminished ability of their bone marrow cells to transfer antibody-forming capacity to sheep red blood cells. The significance of these results, with respect to the origin of the antigen-reactive and antibody-forming cells in the rabbit, is discussed.
FURTHER IMPLICATION OF MURINE LEUKEMIA-LIKE VIRUS IN THE DISORDERS OF NZB MICE
Authors: Mellors Robert C. , Aoki Tadao , Huebner Robert J.
Published: June 2004

Further evidence implicating murine leukemia-like virus in the disorders of NZB mice was afforded by a study of antigens associated with murine leukemia virus (MuLV). MuLV group antigens were prevalent in extracts of spleen, kidney, and, to a lesser extent, thymus throughout a substantial portion of the life span of NZB mice as well as in extracts of lymphomas and sarcomas indigenous to the strain.

G (Gross) soluble antigen, type-specific antigen, was first detected in plasma of untreated NZB mice at 3 months of age. G soluble antigen production increased thereafter in line with age, with 50% of reactions becoming positive at 5.3 months and 100% at 7 to 9 months.

From months 3 to 9, the time-response curve for positive conversion of direct antiglobulin (Coombs) tests in untreated NZB mice corresponded closely to that for G soluble antigen production.

Beyond the 9th month, G soluble antigen underwent elimination from the plasma of NZB mice, with positive reactions reduced to 50% at 13.3 months and to 0% at 18 months. G natural antibody was first detected in the serum of NZB mice at about 10 months of age and increased thereafter in line with age. The curves for G antibody production and G soluble antigen elimination bore a reciprocal relation to each other with crossover at 50% response occurring at 13.3 months.

Significant proteinuria, a functional manifestation of membranous glomerulonephritis, became increasingly prevalent in female NZB mice as G soluble antigen was eliminated from plasma. Cumulative mortality of female NZB mice, mainly attributable to renal glomerular disease, increased in phase with G antibody production. MuLV group antigens were identified in the glomerular lesions by the immunofluorescence method.

Positive conversion of direct antiglobulin tests was significantly delayed by vaccinating baby NZB mice with formaldehyde-inactivated cell-free filtrates of older NZB mouse spleens. The plasmas of vaccinated NZB mice with negative direct antiglobulin reactions at 4 to 7 months were likewise negative when tested for G soluble antigen. The 50% response time for G antibody production in the vaccinated NZB mice occurred at 7.3 months, that is, 6 months earlier than in untreated NZB mice.

The collective findings implicate murine leukemia-like virus in the etiology of autoimmune hemolytic disease and membranous glomerulonephritis, as well as malignant lymphoma, of NZB mice and suggest that virus-specified cell-surface and soluble antigen is a factor in the immunopathogenesis of the renal disease and possibly also the autoimmune hemolytic disease.
ISOLATION OF A FRAGMENT (C3a) OF THE THIRD COMPONENT OF HUMAN COMPLEMENT CONTAINING ANAPHYLATOXIN AND CHEMOTACTIC ACTIVITY AND DESCRIPTION OF AN ANAPHYLATOXIN INACTIVATOR OF HUMAN SERUM
Authors: Bokisch Viktor A. , Müller-Eberhard Hans J. , Cochrane Charles G.
Published: June 2004

A small fragment of C3, called C3a, which has smooth muscle contracting activity, was isolated by three different methods. At pH 8.6, C3a behaved as cation, and using the Archibald method, its mol wt was determined to be 7000. A specific antiserum to C3a showed the fragment to be antigenically distinct from the rest of the C3 molecule, i.e., the C3b portion. The same antiserum and an anti-whole C3 were able to inhibit the biologic activity of C3a. In addition to anaphylatoxin activity, leukocyte chemotactic activity was shown to reside in C3a. Treatment with trypsin caused the cationic fragment to become anionic and abolished the anaphylatoxin but not the chemotactic activity.

C3a fragments with identical biologic activity and comparable cationic properties, as determined by acid disc electrophoresis, were obtained by treatment of C3 with C3 convertase, C3 inactivator complex, trypsin, and plasmin. Thrombin produced a similar C3 fragment which was inactive. It was concluded that C3a corresponds to an unusually basic portion of C3 which may be liberated by attack of a variety of enzymes on a highly susceptible region of the native C3 molecule.

C3b was cleaved by trypsin and less efficiently by thrombin or plasmin into two antigenically distinct pieces: the larger C3c fragment corresponding to ß1A and the smaller C3d fragment to α2D of aged serum. The c- and the d-fragments were separated and characterized.

Isolated C3a rapidly lost its anaphylatoxin activity when treated with small amounts of a partially purified, thermolabile 10S α-pseudoglobulin of human serum. The conditions of inactivation suggested an enzymatic reaction. The anaphylatoxin inactivator also destroyed the activity of C5-derived anaphylatoxin and of lysyl bradykinin.
THE REQUIREMENT OF MORE THAN ONE ANTIGENIC DETERMINANT FOR IMMUNOGENICITY
Authors: Rajewsky K. , Schirrmacher V. , Nase S. , Jerne N. K.
Published: June 2004

Rabbits primarily stimulated with a BSA (bovine serum albumin)-sulfanilic acid complex will produce a good secondary response to the sulfanilic acid hapten if the carrier used in the secondary stimulus is again BSA, and not if the secondary carrier is HGG (human gamma globulin). In the latter situation, a good secondary response is obtained, however, if the rabbits are pretreated a few weeks earlier with free HGG. We conclude that the immune stimulus involves the recognition of carrier determinants unrelated to the hapten.

As the receptors for recognition of unrelated determinants are probably situated on different cells, we suggest that the immune stimulus leading to antibody formation requires the interaction of two antigen-bridged cells.
COMPARISON OF REACTIONS OF ANTIBODIES TO RAT COLLAGEN AND TO RAT KIDNEY IN THE BASEMENT MEMBRANES OF RAT RENAL GLOMERULI
Authors: Rothbard Sidney , Watson Robert F.
Published: June 2004

By in vivo and in vitro methods of immunofluorescence, antibody to rat collagen and to rat kidney show the same regular, linear fluorescence following the outlines of the renal glomerular capillaries. Absorption of each antiserum with its homologous antigen completely removed the antibody for immunofluorescence, while absorption with the heterologous antigen had no effect. The nephrotoxicity persisted in the anti-kidney serum absorbed with collagen.

By pretreatment of frozen normal rat kidney sections with various enzymes followed by immunofluorescence, it was shown that trypsin and hyaluronidase had no effect on the subsequent fluorescence of either antibody; papain reduced the fluorescence; and pepsin and Pronase acted on both antigens so that no fluorescence was present. One preparation of neuraminidase, derived from V. cholerae, reduced fluorescence of both antibodies in some preparations, but the same enzyme derived from influenza virus or C. perfringens had no effect on either. Collagenase completely prevented fluorescence of the antibody to collagen and had no effect on that to rat kidney.

The findings in this study show that the antibody to collagen is directed to collagen in rat renal glomerular basement membranes and that the antibody to rat kidney reacts with some antigen other than collagen in these membranes.
HUMAN IMMUNITY TO THE MENINGOCOCCUS
Authors: Gotschlich Emil C. , Goldschneider Irving , Artenstein Malcolm S.
Published: June 2004

High molecular weight group A and group C meningococcal polysaccharides had no significant toxicity for mice or guinea pigs. Furthermore, these polysaccharide preparations contained negligible amounts of biologically active endotoxin.

The high molecular weight group A and group C meningococcal polysaccharides were excellent immunogens in six human volunteers. Antibodies belonging to the immunoglobulin classes IgG, IgM, and IgA were produced.

These antibodies were highly meningococcocidal in the presence of complement.
PHYSICAL SEPARATION OF HEMOPOIETIC STEM CELLS DIFFERING IN THEIR CAPACITY FOR SELF-RENEWAL
Authors: Worton R. G. , McCulloch E. A. , Till J. E.
Published: June 2004

Bone marrow cells in suspension were separated into a number of fractions on the basis of cell size by sedimentation at unit gravity through gradients of fetal calf serum. The colony forming units (CFU) from the various fractions were tested for their self-renewal capacity using a double transplantation technique. The results indicate that the CFU in the fractions containing slowly sedimenting cells have an increased capacity for self-renewal in comparison with CFU in fractions containing rapidly sedimenting cells. In addition, a culture method was used to select populations containing CFU with increased self-renewal capacity, and these CFU were shown to sediment slowly in comparison with CFU of lower self-renewal capacity obtained from control cultures. It may be concluded that at least part of the heterogeneity observed in the CFU content of individual spleen colonies arises from the composition of the initial cell suspension, probably from intrinsic differences between the stem cells themselves.
THE ROLE OF MYCOBACTERIA AND THE EFFECT OF PROTEOLYTIC DEGRADATION OF THYROGLOBULIN ON THE PRODUCTION OF AUTOIMMUNE THYROIDITIS
Authors: Weigle W. O. , High Gloria J. , Nakamura R. M.
Published: June 2004

Data are presented which suggest that the initial event involved in experimental autoimmune thyroiditis following injection of rabbits with homologous thyroglobulin in complete Freund's adjuvant is alteration of the thyroglobulin. Alteration of the thyroglobulin does not occur during incorporation into the adjuvant or in vitro storage in the adjuvant, and the mycobacteria in the adjuvant have no direct effect on the thyroglobulin. Most likely, the alteration results from an increase in hydrogen ion concentration within cells or local areas in the granuloma and the subsequent action of proteolytic enzymes. These conditions are probably established in the granuloma as the result of neutrophilic response to the mycobacteria in the adjuvant.

Rabbits injected with aqueous preparations of homologous thyroglobulin partially degraded in vitro with pepsin at acid pH produced antibody to native thyroglobulin and developed thyroiditis. Most of these rabbits responded to a subsequent injection of native thyroglobulin given 1 month later.
IMMUNE RESPONSES IN VITRO
Authors: Pierce Carl W.
Published: June 2004

A cell suspension culture system combined with a procedure which separates most macrophages from lymphoid cells was used to investigate some of the cellular requirements for direct and indirect plaque-forming cell responses by nonprimed and primed mouse spleen cells in vitro.

The plaque-forming cell response to heterologous erythrocytes in cultures of nonprimed spleen cells required both macrophages and lymphoid cells for its development. A significant indirect plaque-forming cell response did not develop in cultures of nonprimed spleen cells.

In contrast, cultures of separated or macrophage-poor lymphoid cells from primed mice exhibited increasing responses relative to the response of unseparated spleen cells as the interval after priming increased. The cultures of separated lymphoid cells were not entirely free of phagocytic cells. Despite some evidence which suggests that these phagocytic cells had little function in the response, one cannot ascertain whether the lymphoid cells were responding directly to a second contact with antigen or whether the few contaminating phagocytic cells were performing a function essential to the response by the lymphoid cells. Physiologically different populations of cells appear to develop after priming and are able to respond in vitro in a macrophage-poor culture. Some of the properties of these populations suggest that they are "memory cell" pools containing precursors of direct and indirect plaque-forming cells highly susceptible to a second antigenic stimulus.
THE NATURE OF THE ANTIGENIC DETERMINANT IN A GENETIC CONTROL OF THE ANTIBODY RESPONSE
Authors: Mozes Edna , McDevitt Hugh O. , Jaton Jean-Claude , Sela Michael
Published: June 2004

The response of inbred mouse strains to two polypeptides derived from multichain polyprolines, (T,G)-Pro--L and (Phe,G)-Pro--L, is different from the response of the same mouse strains to a similar series of polymers built on multi-poly-D,L-alanyl--poly-L-lysine, although the same short sequences of amino acids are attached to the side chains of the polypeptides in the two series. These results indicate that a portion of the side chain (e.g. polyalanine or polyproline) participates in the antigenic determinant. This was confirmed by studying the response of different mouse strains to two kinds of polypeptides: (T,G)-Pro-A--L 717 and 718 and (T,G)-A-Pro--L 719 and 721.

Antibody assay of antisera to (Phe,G)-Pro--L with the cross-reacting antigens (T,G)-Pro--L and (Phe,G)-A-L indicates that different inbred mouse strains make antibodies specific for different parts of the same polypeptide. Thus, antibody from DBA/1 mice reacts almost exclusively with the (Phe,G) sequence, while SJL antisera bind only (T,G)-Pro--L and fail to bind (Phe,G)-A-L.

The immune responses to the same amino acids on two different polypeptides (i.e. A--L and Pro--L) appear to be under separate genetic control.
C3 LEUKOTACTIC FACTORS PRODUCED BY A TISSUE PROTEASE
Authors: Hill Jeffrey H. , Ward Peter A.
Published: June 2004

When various rat tissues are incubated in homologous serum, a factor chemotactic in vitro for neutrophils is generated. The amount of chemotactic activity is a function of duration of incubation and the quantity of heart tissue or serum employed. Addition of trypsin inhibitor or antibody to the third component of complement (C3) precludes generation of chemotactic activity. In addition, antibody to C3 ablates chemotactic activity even after its formation.

Purified human C3 (ß1C-globulin) effectively substitutes for serum in the generation of chemotactic activity by heart tissue. The active product, as determined by gel filtration or by ultracentrifugal analysis in a sucrose density gradient, appears to be a cleavage product of C3 with a molecular weight of approximately 14,000. In addition, a larger C3 fragmentation product varying in molecular weight, depending upon experimental conditions, is also found.

The protease in rat heart tissue capable of cleaving C3 into chemotactic fragments is a serine esterase with trypsin-like properties and can be inhibited by organophophorous compounds or trypsin inhibitors. The use of amino acid esters in the manner of competitive substrate inhibition confirms the trypsin-like nature of the protease.

The presence of a protease in heart, and presumably in other normal tissues, capable of fragmenting C3 into factors with chemotactic activities may explain the development of the acute inflammatory response when tissues are non-specifically injured. If true, this would reinforce the role of the complement system in the mediation of nonimmunologically induced inflammation.
LACTOFERRIN, AN IRON-BINBING PROTEIN NI NEUTROPHILIC LEUKOCYTES
Authors: Masson P. L. , Heremans J. F. , Schonne E.
Published: June 2004

Lactoferrin, an iron-binding protein previously shown to occur in many external secretions, is identified as one of the major proteins present in human and guinea pig neutrophilic polymorphonuclear leukocytes.

The identification of this protein in leukocyte extracts was based upon a comparison of its electrophoretic, antigenic, and iron-combining properties with the corresponding properties of the same protein isolated from human and guinea pig milk. Immunochemical quantitations showed that lactoferrin occurs in human neutrophilic leukocytes at the concentration of 3 µg per 106 cells. Tissue cultures from guinea pig bone marrow and spleen actively synthesized the protein, as shown both by net production of lactoferrin and incorporation of labeled amino acids into the protein. Immunohistochemical data indicate that lactoferrin first appears in myeloid cells at the stage of the promyelocyte.
SMALL INTESTINAL MUCOSAL CELL PROLIFERATION AND BACTERIAL FLORA IN THE CONVENTIONALIZATION OF THE GERMFREE MOUSE
Authors: Khoury Kenneth A. , Floch Martin H. , Hersh Theodore
Published: June 2004

The relationship between intestinal colonization and the small bowel mucosal cellular proliferation rate during conventionalization of the germfree mouse was examined. 16 mice were maintained under standard germfree conditions, and 54 others were conventionalized. Migration of the small bowel epithelial cells was followed by radioautography with administration of tritiated thymidine. Colonization was followed by qualitative and quantitative bacteriological fecal analyses. The percentages of the villi labeled (as determined by cell count) 24, 48, and 72 hr following thymidine administration showed immediate progression in the conventionalized animals from the germfree villus migration time (4 days) toward the conventional villus migration time (2 days). The epithelial migration rate of animals conventionalized for 8 days was comparable to that of conventional animals.

After conventionalization, aerobic and anaerobic organisms undergo a period of extensive multiplication; however, 72 hr later the number of these microorganisms cultured in the stool decrease and are similar to those recovered from normal animals. Coliforms and streptococci are recovered in large numbers only in the first days after conventionalization, while the Bacteroides are first recovered in significant numbers on the fifth day of conventionalization. Except for smaller numbers of Bacteroides, the bacterial populations in the stools of the conventionalized animals are qualitatively and quantitatively similar by the eighth day of conventionalization to those of true conventional mice.

Adaptive balance between cell proliferation and sloughing, and thus migration rate, begins immediately after conventionalization of germfree animals as bacterial populations establish themselves throughout the gastrointestinal tract, and results in a doubling of the mucosal cell turnover after 8 days. At this time both the small intestinal epithelial cell migration rate and the intestinal microflora are similar to those of conventional animals.
PHYSICAL, CHEMICAL, AND ULTRASTRUCTURAL STUDIES OF WATER-SOLUBLE HUMAN AMYLOID FIBRILS
Authors: Pras Mordechai , Zucker-Franklin Dorothea , Rimon Abraham , Franklin Edward C.
Published: June 2004

Amyloid fibrils were isolated from the tissues of nine patients with amyloidosis in a state of high purity by homogenization of the tissue followed by extraction with distilled water. Physical, chemical, and ultrastructural studies suggest that amyloid fibrils from different individuals resemble each other, but are not identical. In tissue sections as well as by negative staining of isolated fibrils, morphologic variations were observed. Among the isolated fibrils at least three types were noted. The majority resembled those described previously. However, one subject had two types of fibrils which differed in size and appearance.

Most of the preparations sedimented as a single component with a sedimentation coefficient of 45–50S or as a larger polymer. However, two of the preparations had sedimentation coefficients of 8–9S, and a third one had a major 95S component and a minor 9S fraction.

While the preparations of amyloid were not sufficiently pure for amino acid analyses, peptide maps demonstrated differences among amyloid preparations from different subjects.

The amyloid fibrils in their native state proved to be remarkably resistant to digestion by a number of proteolytic enzymes. Several chemical methods were tried to produce smaller subunits. Of these, the most successful one was the use of 0.1 M NaOH which yielded a smaller, soluble fraction with sedimentation coefficients ranging from 1.1 to 2.8S. Accompanying this degradation, there was little loss of peptides or carbohydrates. Based on the results of the chemical analyses, it is estimated that the subunit produced by sodium hydroxide had a molecular weight of approximately 35,000–40,000.
FLUID PHASE DESTRUCTION OF C2hu BY C1hu
Authors: Gigli Irma , Austen K. Frank
Published: June 2004

It has been demonstrated that C1 isolated in the unactivated form fails to inactivate C4 or C2 in the fluid phase, while the activated molecule, C1 rapidly converts C4 to hemolytically inactive C4i, but does not efficiently inactivate C2. The production and presence of C4i now confers on C1 the ability to rapidly inactivate C2. After heating at 56°C, so as to destroy the hemolytic activity, heat inactivated C1 is still capable of inactivating C4 but the presence of C4i no longer confers an ability to inactivate C2. Studies with the subunits of C1–C1q, C1r, C1s, indicate that the action of C1s on C2 can be inhibited by C1r and that this inhibition is reversed by the presence of homologous C4. These studies indicate that the interaction of C4i with a heat labile receptor conformation in C1 uncovers a masked specificity for C2.
CELLS INVOLVED IN THE IMMUNE RESPONSE
Authors: Abdou Nabih I. , Rose Bram , Richter Maxwell
Published: June 2004

By appropriate irradiation and cell transfer experiments, a direct correlation was observed between the presence of viable and immunologically active antigen-reactive cells and the capacity of the rabbits to respond following immunization. Rabbits given 800 R total body irradiation were unable to elicit a humoral immune response nor did they possess significant numbers of antigen-reactive cells. The ability to respond with humoral antibody formation did not reappear until antigen-reactive cells could be detected. These results strongly indicate that the presence of competent antigen-reactive cells are necessary for the successful induction of the humoral immune response in the rabbit.
QUANTITATIVE INVESTIGATIONS OF IDIOTYPIC ANTIBODIES
Authors: Daugharty Harry , Hopper John E. , MacDonald A. Bruce , Nisonoff Alfred
Published: June 2004

Specifically purified anti-p-azobenzoate antibodies of the IgG class from individual rabbits were used to elicit anti-idiotypic antibodies in recipient rabbits. Allotypes of each donor and recipient were matched. When polymerized antibodies were used for immunization, more than 80% of the recipients responded with the formation of antibodies that precipitated the monomeric donor antibody. Percentages of precipitable molecules in the donor antibody population (D) varied from 4 to 56. As little as 4% was readily detectable by the Ouchterlony method or precipitin test. Specificity of the reaction was tested by double diffusion in agar gel against a panel of purified antibenzoate antibodies from 14 heterologous rabbits and, quantitatively, in three systems by measurement of the extent of coprecipitation of heterologous, radiolabeled antibenzoate antibodies. No cross-reactions were observed. Reactions were shown to be attributable to antibenzoate antibodies in the donor serum, and contributions of allotypic reactions were excluded. In three systems investigated quantitatively, and in one studied qualitatively, two recipients of the same donor antibody produced anti-antibody that reacted with essentially the same subfraction of the donor antibody population. The findings that only a portion of the D population is immunogenic, and that the same subfraction is frequently immunogenic in different recipients, suggest that the immunogenic population comprises a limited number of homogeneous groups of antibody molecules. This is supported by the small number of bands usually observed by the Ouchterlony technique.

Quantitative methods of analysis should provide an approach to the study of cell populations producing antibodies of a particular idiotype.
BINDING BETWEEN COMPONENTS OF THE TUBERCLE BACILLUS AND HUMORAL ANTIBODIES
Authors: Minden Percy , Farr Richard S.
Published: June 2004

Studies were undertaken to find a substance or substances for use in primary binding types of tests to detect humoral antibodies in rabbits and monkeys exposed to the tubercle bacillus that would distinguish between strains of mycobacteria. The antigen employed was a component of the 5159 strain of Mycobacterium tuberculosis that was obtained following sonification, ultracentrifugation, electrophoresis, and elution from a preparative polyacrylamide column. When the antigen was labeled with 131I, specific binding was observed in sera from immunized rabbits and BCG-protected rhesus monkeys by the radio-gel electrophoresis and ammonium sulfate tests.

This component was partially characterized, and its major antigenic determinants were associated with anionic mucopolysaccharides. Electrophoretically at pH 8.3 it migrated anodally to albumin, its molecular weight was between 9,000 and 12,000, and it was soluble in 50% saturated ammonium sulfate. Binding to antibody was destroyed after treatment with Pronase, but not after DNase or RNase.

Inhibition of the reaction, as measured by the ammonium sulfate test, between the 131I-labeled component and antisera from rabbits that had been immunized with sonicated 5159 organisms, was studied. These experiments demonstrated a capacity to define subtle similarities and differences among different mycobacteria and mycobacterial components. Some microorganisms not clearly related to mycobacteria also partially inhibited this reaction, suggesting that they shared antigenic groups with the component derived from 5159 organisms. The studies described suggested the advisability of using direct primary tests and purified components of mycobacteria to differentiate further the antigenic groups between individual pathogenic mycobacteria and between pathogenic and nonpathogenic organisms.
LASTING BIOLOGICAL EFFECTS OF EARLY ENVIRONMENTAL INFLUENCES
Authors: Lee Chi-Jen , Dubos Rene
Published: June 2004

Physicochemical and immunological techniques have been used in an attempt to characterize a filterable agent, separated from the intestines of mice raised under ordinary conditions of husbandry, which produces a lasting depression of weight in specific pathogen-free (SPF) mice when administered to them orally shortly after birth. Although this agent has not yet been identified, it will be tentatively designated here as enterovirus.

The mouse enterovirus can be readily sedimented by ultracentrifugation and by precipitation at pH 4.3; it does not pass through cellophane membranes. Its infective power is completely destroyed by ultraviolet radiation, but is resistant to heating at 56°C, exposure to ether, treatment with trypsin, ribonuclease, and deoxyribonuclease.

Dialysis and treatment with ether and nucleases greatly increase the infective activity of the intestinal filtrates containing the enterovirus, a finding which suggests that these procedures eliminate or destroy some inhibitory substance(s).

The mouse enterovirus causes hemagglutination of mouse red blood cells. When injected into rabbits, it elicits in them an immune response that renders their serum capable of neutralizing its weight-depressing activity. As measured by inhibition of hemagglutination or complement fixation, the sera of infected mice do not exhibit any significant activity against usual mouse viruses.

Centrifugation of the mouse enterovirus in 50%–20% sucrose gradient gave almost complete recovery of the infectivity and of hemagglutinating activity in the same fraction. In contrast, the protein content of the material was distributed through the various fractions. Consequently, this procedure resulted in a marked increase of specific activity.
GENETIC INFLUENCE ON THE DEVELOPMENT OF RENOPRIVAL HYPERTENSION IN PARABIOTIC RATS
Authors: Knudsen K. D. , Iwai J. , Heine M. , Leitl G. , Dahl L. K.
Published: June 2004

Rats from two strains with opposite constitutional predisposition to hypertension were joined in parabiosis and one partner was nephrectomized. The influence of genetic factors and of diet on the blood pressures of the two classes of parabionts, operated and intact, indicated that renoprival hypertension occurred with equal frequency in rats from both strains; that the development of renoprival hypertension depended on the influence from an intact S partner, or on a high salt intake, or on both.

A nephrectomized S rat developing renoprival hypertension did not induce high blood pressure in its intact R partner. In this respect renoprival hypertension differs from salt and renal hypertension.

The findings are interpreted to mean that the hypertensinogenic agent specific for S rats is produced by S kidneys.
IMMUNOLOGICAL EFFECTS OF THERMAL INJURY
Authors: Rapaport Felix T. , Sampath Angus , Kano Kyiochi , McCluskey Robert T. , Milgrom Felix
Published: June 2004

Unilateral thermal injury to guinea pig testes has elicited the formation of organ- and species-specific antibodies against a testicular antigen. The antibodies were of an IgG1 nature, and reacted with autologous as well as homologous testicular extracts.

A significant number of burned animals showed pathological changes in the contralateral testis, which were similar to those observed after the induction of experimental allergic orchitis by active immunization with testicular tissue.

These results indicate that thermal injury may be associated with autoimmunization of the host by the injured organ.
IN VITRO INDUCTION OF SPECIFIC UNRESPONSIVENESS OF IMMUNOLOGICALLY REACTIVE, NORMAL BONE MARROW CELLS
Authors: Singhal Sharwan K. , Wigzell Hans
Published: June 2004

Normal rabbit bone marrow cells have been studied according to their immunological reactivity in vitro. The test system involved stimulation by antigen after the subsequent stimulation into cellular proliferation by measuring the uptake of tritium-labeled thymidine. Specific separation of immunological reactivity was obtained by filtration of cells through antigen-coated bead columns. All experimental evidence supported the view that this separation was due to the existence of preformed antibody molecules on the outer cell surface of the antigen-recognizing cells.

The response to antigenic stimulation was shown to be strictly dose related and, using supraoptimal concentrations of one antigen, no increased DNA synthesis was recorded. That this state of unresponsiveness represented a state of immunological paralysis was indicated by the normal response of these cells to stimulation by a second antigen in optimal concentration.

Thus both methods, cell separation on antigen-coated columns or induction of specific unresponsiveness by antigen in vitro, can produce a cell population specifically devoid of cells reactive against a given antigen.
RELEASE OF VASOACTIVE AMINES FROM RABBIT PLATELETS INDUCED BY SENSITIZED MONONUCLEAR LEUKOCYTES AND ANTIGEN
Authors: Henson P. M.
Published: June 2004

The immunological release of vasoactive amines from rabbit platelets by a mechanism requiring blood leukocytes has been described. The reaction involved leukocytes from an immunized rabbit, antigen, and platelets and did not require the complement system. The leukocytes appeared to be mononuclear, had the ability to adhere to glass, and were found in greater numbers in the blood than in spleen, lymph nodes, thoracic duct lymph, bone marrow, and peritoneal cavity washings.

Part or all of the effect on the platelets appeared to result from active release of a soluble factor from the leukocytes, which induced vasoactive amine release from the platelets. The platelets were not lysed during this reaction and the release of vasoactive amines required the active participation of metabolic pathways in the platelet.
GENETIC INFLUENCE ON THE RENIN-ANGIOTENSIN SYSTEM
Authors: Iwai Junichi , Knudsen Knud D. , Dahl Lewis K.
Published: June 2004

Two strains of rats with opposite genetic propensity for hypertension were tested for: (a) the sensitivity to injections of angiotensin and renin, and (b) the influence of their plasma on the reaction velocity of renin and its substrate in vitro.

Intact hypertension-prone (S) rats on low salt had higher sensitivity to angiotensin and a lower sensitivity to renin than hypertension-resistant (R) rats. High NaCl diet did not change the response of the R rats to these injections, but increased the response to renin and angiotensin in intact S rats. Bilateral nephrectomy caused increased response to renin and a decreased response to angiotensin in the S rats, so that both strains were equivalent after bilateral nephrectomy. In vitro, plasma from intact S rats inhibited the activity of hog renin. Plasma from R rats showed no inhibition. The inhibitor disappeared after bilateral nephrectomy. It was speculated that renin inhibitor may be involved in the development of hypertension by increasing sensitivity to angiotensin and other hypertensinogenic stimuli.
IN VITRO INTERACTION OF MOUSE HEPATITIS VIRUS AND MACROPHAGES FROM GENETICALLY RESISTANT MICE
Authors: Shif Ilan , Bang Frederik B.
Published: June 2004

Peritoneal macrophages from genetically resistant C3H mice and genetically susceptible Princeton (PRI) mice adsorbed the MHV (PRI) strain of mouse hepatitis virus equally well. The difference between the permissive cells and the nonpermissive ones seems to reside in the ability of the former to "eclipse" the virus and, subsequently, support virus replication. C3H cells exposed to low multiplicities of the virus remained intact with no demonstrable viral replication. Virus, taken up by the resistant cells, was protected from heat and underwent slow inactivation while few or no virus particles were released into the medium.