Infectious Bronchitis Virus Immune Responses in the Harderian Gland upon Initial Vaccination.

In recent years, Arkansas Delmarva Poultry Industry (ArkDPI)-derived infectious bronchitis (IB) virus (IBV) vaccines have been used to characterize the immune responses of chickens subsequent to vaccination on day of hatch or beyond. Perhaps because ArkDPI vaccines display increased heterogeneity, the results on cell immune responses have shown ambiguity. In the current study, we investigated the effects of vaccination with a highly stable and homogeneous Massachusetts (Mass)-type vaccine on days 1 or 7 of age on Harderian gland (HG) responses. Confirming previous studies, both IBV serum antibodies and lachrymal IgA levels were greater upon vaccination on day 7 compared with vaccination on day 1 of age. Unlike results with ArkDPI viruses, a clear trend was detected for both B and T cells in the HG after Mass-type vaccination. Consistent with antibody responses, B- and T-helper (CD3+CD4+) cell frequencies were higher in birds vaccinated on day 7 of age. Cytotoxic T cells (CD3+CD8+) were also increased compared with chickens vaccinated on day 1 of age. Depending on the most likely age of IB outbreaks to occur in a particular region, postponing the first IBV vaccination may optimize immune responses.

Nota de Investigación- Respuestas inmunes al virus de la bronquitis infecciosa en la glándula de Harder después de una vacunación inicial En los últimos años, las vacunas contra el virus de la bronquitis infecciosa (IB) derivadas de la cepa Arkansas Industria Avícola de Delmarva (ArkDPI) se han utilizado para caracterizar las respuestas inmunes de los pollos después de la vacunación en el día de la eclosión o en días posteriores. Posiblemente debido a que las vacunas ArkDPI muestran una mayor heterogeneidad, los resultados sobre las respuestas inmunes celulares han mostrado ambigüedad. En el presente estudio, se investigaron los efectos de la vacunación con una vacuna de tipo Massachusetts (Mass) que es altamente estable y homogénea en los días uno o siete de edad con relación a las respuestas en la glándula de Harder (HG). Confirmando estudios previos, tanto los anticuerpos séricos contra el virus de la bronquitis infecciosa como los niveles de IgA lacrimal fueron mayores tras la vacunación al día siete en comparación con la vacunación al primer día de edad. A diferencia de los resultados con los virus ArkDPI, se detectó una clara tendencia para las células B y T en la glándula de Harder después de la vacunación con el tipo Mass. De acuerdo con las respuestas de anticuerpos, las frecuencias de células B y T cooperadoras (CD3+ CD4+) fueron más altas en las aves vacunadas en el día siete de edad. Las células T citotóxicas (CD3+CD8+) también aumentaron en comparación con los pollos vacunados en el primer día de edad. Dependiendo de la edad más probable en que ocurran brotes por bronquitis infecciosa en una región en particular, posponer la primera vacuna contra el IBV puede optimizar las respuestas inmunes.

Novel insights into the host immune response of chicken Harderian gland tissue during Newcastle disease virus infection and heat treatment.

BACKGROUND: Newcastle disease virus, in its most pathogenic form, threatens the livelihood of rural poultry farmers where there is a limited infrastructure and service for vaccinations to prevent outbreaks of the virus. Previously reported studies on the host response to Newcastle disease in chickens have not examined the disease under abiotic stressors, such as heat, which commonly experienced by chickens in regions such as Africa. The objective of this study was to elucidate the underlying biological mechanisms that contribute to disease resistance in chickens to the Newcastle disease virus while under the effects of heat stress. RESULTS: Differential gene expression analysis identified genes differentially expressed between treated and non-treated birds across three time points (2, 6, and 10 days post-infection) in Fayoumi and Leghorn birds. Across the three time points, Fayoumi had very few genes differentially expressed between treated and non-treated groups at 2 and 6 days post-infection. However, 202 genes were differentially expressed at 10 days post-infection. Alternatively, Leghorn had very few genes differentially expressed at 2 and 10 days post-infection but had 167 differentially expressed genes at 6 days post-infection. Very few differentially expressed genes were shared between the two genetic lines, and pathway analysis found unique signaling pathways specific to each genetic line. Fayoumi had significantly lower viral load, higher viral clearance, higher anti-NDV antibody levels, and fewer viral transcripts detected compared to Leghorns. Fayoumis activated immune related pathways including SAPK/JNK and p38 MAPK signaling pathways at earlier time points, while Leghorn would activate these same pathways at a later time. Further analysis revealed activation of the GP6 signaling pathway that may be responsible for the susceptible Leghorn response. CONCLUSIONS: The findings in this study confirmed our hypothesis that the Fayoumi line was more resistant to Newcastle disease virus infection compared to the Leghorn line. Within line and interaction analysis demonstrated substantial differences in response patterns between the two genetic lines that was not observed from the within line contrasts. This study has provided novel insights into the transcriptome response of the Harderian gland tissue during Newcastle disease virus infection while under heat stress utilizing a unique resistant and susceptible model.

Class B CpG ODN stimulation upregulates expression of TLR21 and IFN-γ in chicken Harderian gland cells.

This study aimed to evaluate the response of Harderian gland (HG) cells after in vitro stimulation with class B synthetic oligodeoxyribonucleotides (ODN) containing CpG motifs. This knowledge is of importance for the development of mucosal vaccines for poultry, such as eye-drop or spray vaccines, to determine if class B CpG ODN can act as an vaccine adjuvant or as a prophylactic treatment mainly against respiratory disease viruses. The relative expression of Toll-like receptor 21 (TLR21), interferon (IFN)-γ, interleukin (IL)-1ß and IL-10 genes were quantified at 1, 3, 6 and 18 h post-stimulation of HG cells from 5-week-old birds. In addition, it was also investigated if expression of these genes was affected by the age of the birds (differences between 5- and 12-week-old birds), concentrations of ODN or cell preparation method used. Class B CpG ODN induced upregulation of TLR21 and IFN-γ mRNA expression levels at 1h post-stimulation depending on concentration of ODN used but only in HG cells isolated from young birds.

Mobilization of immunoglobulin (Ig)-containing plasma cells in Harderian gland, cecal tonsil and trachea of broilers vaccinated with Newcastle Disease Vaccine.

Immunohistochemical studies of Harderian gland, cecal tonsil and trachea of various groups of broiler chickens and the response of Baby Chick Ranikhet Disease Vaccines (BCRDV) on the mobilization of Igs-cells during postnatal development of organs was investigated in the Dept. of Anatomy and Histology, Bangladesh Agricultural University, Mymensingh. In this study twelve chickens were grouped into vaccinated broilers (D14 and D28) which had received vaccines first at D3 of age and a booster dose given at D13; and non-vaccinated broilers (D1) which had not been vaccinated. In this study, it was observed that the frequency and distribution of Igs-positive cells were higher at D14 and at D28 rather than D1. Among Igs-positive cells, the IgG-positive cells were significantly higher than IgM and IgA-positive cells in the Harderian gland of D14 and D28 groups of chickens, however, in day-old chickens, the frequency of IgM-positive cells in this gland were greater. In the cecal tonsil, the frequency and distribution of IgG-positive cells were significantly higher than IgA- and IgM-positive cells both at D14 and D28 ages of chicken. On the other hand, in day-old chickens, the frequency and distribution of IgA-positive cells were insignificantly greater, followed by IgM and IgG-positive cells. In the trachea, few immunoglobulin-containing plasma cells were distributed in the subepithelial layer. IgM-positive cells were higher followed by IgG and IgA-positive cells in the trachea in D14 and D28 groups of chickens. In the same organ, IgG-positive plasma cells were greater than IgA and IgM-positive cells at one-day old. When the data for Harderian gland, cecal tonsil and trachea were compared statistically, it was observed that Igs-positive cells were statistically more common in cecal tonsils in day old chickens, and with the advancement of age, Igs-positive cells were found more in the Harderian gland. In conclusion, with the advancement of age in chickens the Harderian gland uptake is a function of the cecal tonsil due to its functional atrophy.

Local and systemic immune responses following infection of broiler-type chickens with avian Metapneumovirus subtypes A and B.

Infections with avian Metapneumovirus (aMPV) are often associated with swollen head syndrome in meat type chickens. Previous studies in turkeys have demonstrated that local humoral and cell-mediated immunity plays a role in aMPV-infection. Previous experimental and field observations indicated that the susceptibility of broilers and their immune reactions to aMPV may differ from turkeys. In the presented study local and systemic immune reactions of broilers were investigated after experimental infections with subtypes A and B aMPV of turkey origin. Both virus subtypes induced a mild respiratory disease. The recovery from respiratory signs correlated with the induction of local and systemic aMPV virus-neutralizing antibodies, which began to rise at 6 days post infection (dpi), when the peak of clinical signs was observed. In a different manner to the virus neutralizing (VN) and IgG-ELISA serum antibody titres, which showed high levels until the end of the experiments between 24 and 28 dpi, the specific IgA-ELISA and VN-antibody levels in tracheal washes decreased by 10 and 14 dpi, respectively, which may explain the recurring aMPV-infections in the field. Ex vivo cultured spleen cells from aMPV-infected broilers released at 3 and 6 dpi higher levels of IFN-γ after stimulation with Concanavalin A as compared to virus-free birds. In agreement with studies in turkeys, aMPV-infected broilers showed a clear CD4+ T cell accumulation in the Harderian gland (HG) at 6 dpi (P<0.05). In contrast to other investigations in turkeys aMPV-infected broilers showed an increase in the number of CD8alpha+ cells at 6 dpi compared to virus-free birds (P<0.05). The numbers of local B cells in the Harderian gland were not affected by the infection. Both aMPV A and B induced up-regulation of interferon (IFN)-γ mRNA-expression in the nasal turbinates, while in the Harderian gland only aMPV-A induced enhanced IFN-γ expression at 3 dpi. The differences in systemic and local T cell and possibly natural killer cell activity in the HG between turkeys and chickens may explain the differences in aMPV-pathogenesis between these two species.

Production of house finch (Carpodacus mexicanus) IgA specific anti-sera and its application in immunohistochemistry and in ELISA for detection of Mycoplasma gallisepticum-specific IgA.

The IgA antibody response plays a vital role in mucosal immunity because it functions to neutralize pathogens at the mucosal surface and thus impedes attachment to underlying tissues. Although the importance of IgA in the mucosal immunity of galliform birds has been established, studies examining IgA-based immunity in passerine birds are lacking, perhaps due in part to the absence of reagents that can detect passerine IgA. A 469 base pair region of the house finch (Carpodacus mexicanus) IgA heavy chain was PCR-amplified from spleen cDNA and sequenced. The predicted amino acid sequence was found to share 55% and 46% identity with the IgA heavy chain of mallard (Anas platyrhynchos) and chicken (Gallus gallus), respectively. The heavy chain fragment was produced using a bacterial expression system and purified. Rabbit anti-sera were generated against the recombinant protein. The anti-sera reacted with a single house finch serum protein ( approximately 50-55kDa) in Western blot. The anti-sera were used to identify plasma cells in the Harderian gland and conjunctiva of house finches with conjunctivitis associated with Mycoplasma gallisepticum infection. The anti-sera were also utilized in an ELISA to detect M. gallisepticum-specific IgA antibodies in lachrymal samples of infected finches.

Induction of mucosal immunity in the avian Harderian gland with a replication-deficient Ad5 vector expressing avian influenza H5 hemagglutinin.

The chicken Harderian gland (HG) plays an important role in adaptive immune responses upon ocular exposure to avian pathogens such as avian influenza (AI). To determine the role of HGs in generating immunity, chickens were immunized ocularly with an adenovirus (Ad5) vector expressing the AI hemagglutinin H5 gene. The Ad5-H5 vector induced H5 transgene expression and induced H5- and Ad5-specific IgA and IgG spot-forming cells (SFCs) in the HGs. The IgA and IgG SFC peaked on day 9 forAd5 and day 11 for the H5 protein. In addition, Ad5- and H5-specific antibodies were induced in serum. IgA in chicken tears was predominantly dimeric, while in serum monomeric IgA was most abundant. Analysis of HG mRNA confirmed expression of the polymeric immunoglobulin receptor (plgR). These data demonstrated the importance of HGs to generate mucosal and systemic immunity to AI following ocular Ad5-H5 administration to chickens.

Infectious bronchitis virus in the chicken Harderian gland and lachrymal fluid: viral load, infectivity, immune cell responses, and effects of viral immunodeficiency.

We compared detection of infectious bronchitis virus (IBV) by quantitative RT-PCR (qRT-PCR) in tears and trachea of IBV-infected chickens and found that quantitative detection of IBV RNA in tears is more sensitive than in tracheal homogenates. Furthermore, we demonstrated that IBV contained in chicken lachrymal fluid is infectious and that tears of IBV-infected chickens can be used to infect naive chickens. We compared the immune responses to IBV in the Harderian gland and cecal tonsils of immunocompetent chickens and chickens infected with chicken anemia virus (CAV) and/or infectious bursal disease virus (IBDV). Flow cytometry analyses of lymphocytes in Harderian glands and cecal tonsils indicated that the relative abundance of IgM+ B cells in the Harderian glands and cecal tonsils following exposure to IBV in combination with immunosuppressive viruses was reduced compared to chickens infected with IBV alone. CAV, but not IBDV, reduced the CD4+/CD8+ T cell ratios compared to chickens infected with IBV alone. Enzyme-linked immuno-spot forming assays on cells in the Harderian glands and cecal tonsils of IBV-infected chickens indicated that maximum IBV-specific IgA-secreting cell responses were reduced in chickens infected with CAV. IBDV co-infected chickens displayed a delayed IgA response to IBV. Thus immunosuppressive viruses reduced B cells and T helper cells in the Harderian glands and cecal tonsils in response to IBV, and slowed the kinetics and/or reduced the magnitude of the mucosal immune response against IBV. We have shown for the first time that CAV affects pathogen-specific B cell responses in a mucosal effector site.

Immunoglobulin (Ig)-containing plasma cells in the Harderian gland in broiler and native chickens of Bangladesh.

The distribution and frequency of immunoglobulin (Ig)-containing plasma cells, their variations due to sex, and the mode of secretion of Ig cells into the duct system of the Harderian gland was investigated in broiler and native chickens of both sexes in Bangladesh. The Harderian gland is covered by a capsule, and the connective tissue septa divide the gland into numerous unequal-sized numerous lobes and lobules. The Ig-containing plasma cells were located in the interstitial space, interacinar space, apical part of the lobule, and lumina of the lobules of the Harderian gland in both broiler and native chickens. The population of these Ig-containing plasma cells varied in between broiler and native chickens, and also between male and female broiler and native chickens. In the broiler, the number of IgM-containing plasma cells was higher; in contrast, in the native chickens, the population of IgA-containing plasma cells was larger. In the broiler, there were more IgA- and IgG-containing plasma cells in the male; in contrast, there were more IgM-containing plasma cells in female. In native chickens the frequency of IgA-containing plasma cells was greater in the female than male. When the data for broiler and native birds were compared, it was found that there were significantly more IgA- and IgG-containing plasma cells in the native male and female chickens than in the broiler males and females. The secretory Igs were located in the lumina of acini and the duct system of the Harderian gland. In the present study Ig-containing plasma cells were observed to be released in the lumina of the lobules of Harderian gland by the breakdown of acinar tissues in broilers, and by holocrine mode of secretion in the native chicken. These results suggested that the Harderian gland, even though it is not a lymphoid organ as a whole, but acts as an immunopotent organ in chickens, and that the gland in native chicken contains more Ig-containing plasma cells due to their scavenging.

Expression of different classes of immunoglobulin in intraepithelial plasma cells of the Harderian gland of domestic ducks Anas platyrhynchos.

The Harderian gland of chickens contains numerous plasma cells and is considered as a peripheral lymphoid organ. Data about this gland in other avian species are scarce or inexistent. Considering that ducks show some unique characteristics regarding the immune system, which are important in evolutionary context, and that unusual location of plasma cells into the epithelium was recently described in primitive avian species, here we investigated the occurrence and characterized intraepithelial plasma cells in the Harderian gland of ducks, according to the immunoglobulin produced. Numerous intraepithelial plasma cells were found confined to the Harderian gland ducts. Plasma cells were also found in the ducts lamina propria. IgM-positive cells were the most abundant into the epithelium. In contrast, IgY- or IgA-positive cells were predominant in the lamina propria. The constancy of intraepithelial plasma cells in all specimens examined indicates that they may be essential mediator for an effective immunesurvaillance of the ocular mucosa.

Immunohistochemical localization of three different immunoglobulin classes in the Harderian gland of young chickens.

The chicken Harderian gland (HG) was investigated using single immunohistochemical staining for one of the three different immunoglobulins (Igs) followed by Alcian blue/periodic acid Schiff (AB/PAS) staining and triple immunohistochemical staining for all of the Igs with hot water treatment. In the HG of 5-week-old chickens, IgG-containing plasma cells were more frequent than IgA- and IgM-containing cells. These numerous IgG-containing cells were predominantly accumulated in the central region of the stroma, whereas a small number of IgA- and IgM-containing cells were scattered in the peripheral region of the stroma. Also, the plasma cells containing PAS-positive Russell bodies (RBs) exhibited distinct immunoreactivity for one of the Igs, being inversely proportional to the intensity of PAS reaction. The RB-containing cells positive for IgA were more frequent than those positive for IgM, whereas those positive for IgG were very rare. The most distinct feature of the IgG-containing plasma cells was a PAS-positive globule located close to the nucleus. Triple immunostaining with hot water treatment simultaneously identified these three Igs in normal plasma cells and RB-containing ones in the stroma of the chicken HG.

La glándula de Harder: un transductor neuroinmunoendrocrino / The Harderian gland: neuroinmunoendrocrine transductor

La glándula de Harder es una glándula túbulo-alveolar localizada en la parte posterior de la órbita ocular de animales que poseen membrana nictitante. En estos mamiferos la glándula contiene una gran cantidad de lípidos. La glándula de Harder de roedores contiene un pigmento café rojizo, el cual ha sido identificado como porfirina. Las funciones de la glándula de Harder son; síntesis y liberación de ferhormonas, fotoprotección y termorregulación, osmoprotección y se le ha propuesto además un papel inmunoendocrino

Possible migration of harderian gland immunoglobulin A bearing lymphocytes into the caecal tonsil in chickens.

This study was designed to evaluate the significance of the Harderian gland (HG) in the chicken IgA-production system by investigating the influence of HG on the increase of surface IgA (sIgA)-positive cells in other lymphoid organs and also the possibility of migration of HG lymphocytes into other organs. Non-contact culture of splenic or caecal tonsil (CT) lymphocytes with HG whole cells did not enhance the expression of sIgA. The experiment of HG lymphocyte transfer showed that the transferred HG lymphocytes migrated mainly into the CT in the 3-week-old recipient, and mainly into the CT and bursa of Fabricius (BF) in the 6-week-old recipient. It was also found that the transferred sIgA-positive HG lymphocytes migrated selectively into CT in both 3-week-old and 6-week-old recipients. These results indicated that chicken HG plays a central role not only in governing the local immunity in the eyes and respiratory tract but also as an organ which supplies precursory IgA-producing cells involved in local immunity in the intestine.

Phenotypic analysis of resident lymphoid cells in the conjunctiva and adnexal tissues of rat.

The conjunctival associated lymphoid tissue is considered to be an integral part of the mucosal immune system. Under normal circumstances immune mechanisms in mucosal associated lymphoid tissue of the gut and bronchus can selectively suppress, rather than enhance, immune responsiveness to encountered antigens, inducing a state of tolerance. It is possible that conjunctival associated lymphoid tissue can also induce a state of tolerance to encountered antigens. Such a response may be exploited to modulate immune mediated ocular disease. Enhanced tolerance may protect the host against foreign antigen. Alternatively, under certain circumstances when the normal immune system is altered or disrupted the mucosal tissue may act to induce sensitisation and trigger immune mediated disease. The rat is frequently used as an animal model of immune mediated eye disease, but the normal profile of immune cells in the rat conjunctiva has not been studied. This information is essential for meaningful interpretation in the experimental situation. In this study we examined the immunophenotype of lymphoid tissue associated with the conjunctiva, lacrimal gland and Harderian gland of the Lewis rat. CD4+, Ia+ and the monocyte/macrophage population of cells were found predominantly in the substantia propria of the conjuctiva and interstitial connective tissue of the glands. CD8+ cells were distributed mainly in relation to the conjunctival and glandular epithelium. Goblet cells stained strongly with the monoclonal antibody (MAb) MRC OX-39, which is a marker for IL-2 receptors. The overall pattern of distribution of immunocompetent cells in the rat was found to be similar to that reported in humans.

The effects of cyclosporin A and cyclophosphamide on the populations of B and T cells and virus in the Harderian gland of chickens vaccinated with the Hitchner B1 strain of Newcastle disease virus.

The cellular response to conjunctival vaccination with the Hitchner B1 strain of Newcastle disease virus was studied in the Harderian gland (HG) by immunohistochemistry. Bu-1+ cells and all subpopulations of T cells, (CD3+, CD4+, CD8+, TCR gamma delta, TCR alpha beta 1, and TCR alpha beta 2) were in the interstitial tissue between the ducts and the acini. Plasma cells with cytoplasmic IgM were more dispersed than the other cells and outlined the acini. Bu-1+ cells and all subpopulations of T cells increased at least three-fold after vaccination when compared to uninfected birds on the basis of the average cell counts in sections taken at 3, 5, 7, 10, 14, and 20 days after vaccination. The most marked increase was in the CD8+ cells which increased six-fold. Virus replicated for 10 days in cyclophosphamide (Cy) treated birds and for 7 days in cyclosporin A (CsA) treated birds compared with 5 days in untreated birds. Cy treatment prevented an antibody response to NDV and reduced Bu-1+ and IgM cells in the HG by 20-fold. Cy treatment resulted in a doubling of the number of T cells in the HG but these T cells may have been transiently disabled because it also caused a poor response of the lymphocytes in whole blood to the T cell mitogen concanavalin A (ConA). CsA reduced the T cell numbers in the HG and whole-blood responses to ConA by about 4-fold but T cell numbers rebounded to normal resting values after vaccination with NDV. The clearance time was prolonged either by T cells being less numerous than normal after CsA or being disabled after Cy. T cells, but not B cells, may therefore be essential for virus clearance. CD8+ cells expanded more than CD4+ cells after the vaccination of untreated and CsA-treated birds indicating that CD8+ cells may be key players in vaccinal immunity to NDV.

Duck lymphoid organs: their contribution to the ontogeny of IgM and IgY.

The occurrence of mRNAs encoding mu, nu and nu(delta Fc) immunoglobulin heavy chains and lambda light chains in organs of duck embryos from 16 days of incubation and ducklings up to 74 days of age was assessed by Northern hybridization. The mu message was first detected in bursa of Fabricius and spleen at 16 days of incubation and in cervical lymph nodes at 23 days of incubation, but in other organs (bone marrow, buffy coat, Harderian gland, liver) not until 7 17 days after hatching; in general, the appearance of the lambda message paralleled that of mu. Messenger RNAs encoding one or both of the nu isoforms were first detected in cervical lymph nodes at 25 days of incubation, in spleen and bursa in 1-day-old ducklings, in Harderian gland, bone marrow and liver from 10 to 17 days post-hatching and in buffy coat from 46 days. In most organs, the nu(delta Fc) message was detected prior to the nu message and predominated during the experiment; Harderian gland expressed the nu(delta Fc) message exclusively. These results indicate that bursa of Fabricius, spleen and cervical lymph nodes play early roles in the development of B cells and the ontogeny of duck immunoglobulins while other lymphoid organs support the later differentiation of plasma cells, and that IgY and IgY(delta Fc) are probably not simultaneous products of the same plasma cells.

The harderian gland in autoimmune diabetes of the nonobese diabetic mouse.

Infiltration of the nonobese diabetic (NOD) mouse's Harderian gland (HG) was studied in 1-30-week-old animals. A mononuclear cell invasion of this gland is first seen in 8-week-old female mice (i.e., at a slightly later age than that for the onset of infiltration of pancreatic islets). Infiltrating elements are mainly located at the hilus of the gland or at one or two foci (periacinar infiltration) within the parenchyma. In the latter case, a few elements infiltrate the fibrous connective tissue surrounding the acini (one or more) without damaging them. The most severe histopathological lesion was observed in 16-week-old animals; at this time infiltration ranges from a still focal lesion to complete acinar destruction of the gland. Ultrastructural observations confirm that in several cases acinar cells are destroyed and the HG parenchyma is substituted with infiltrating elements, fibroblasts, and connective tissue. HG infiltration is comparable to the pancreatic inflammatory infiltration; the two processes are very similar, though insulitis starts slightly earlier than HG infiltration. Furthermore, as for insulitis and diabetes incidence, HG infiltration affects NOD males less than females. Moreover, immunocytochemistry has shown that T lymphocytes are the prevalent infiltrating element both in pancreatic islets and HGs. Further studies are required to understand the reasons for autoimmune destruction of this gland.

Immune cell proliferation in the harderian gland: an avian model.

Experimentation has been carried out to study proliferation of plasma cells in the chicken Harderian gland (HG) and to determine if a HG factor influences immune cell (i.e., B cell) proliferation. In young chickens, flow cytometric analysis of propidium iodide (PI)-stained plasma cells revealed that the percentages of cells in both the synthetic (S) and mitotic (G2M) phases of the cell cycle were highest between 6 and 9 weeks of age. A pattern of plasma cell depletion and repopulation in the HG was observed following administration of emetine dihydrochloride. At 3 and 5 days posttreatment the plasma cell population decreased, and by 7 days posttreatment repopulation of the gland with plasma cells occurred. This repopulation appeared as a result of plasma cell proliferation within the HG. Anti-5-bromo-2'-deoxyuridine (BrdUrd) staining of frozen sections showed that the numbers of plasma cells incorporating BrdUrd were low at 3 days posttreatment but were as high, or higher than, controls at 5 and 7 days posttreatment. These results were verified with flow cytometric data of PI-stained plasma cells. Data from bursal cell bioassays revealed proliferative activity influenced by a HG factor. Coculture of bursal cells with phorbol dibutyrate and diluted HG supernatants resulted in prolonged and increased proliferation of these cells. It is possible that the HG of chickens supports plasma cell proliferation through the elaboration of a factor which acts like a lymphokine.

The lymphoid substance of the chicken's harderian gland is organized in two histologically distinct compartments.

Light and electron microscopical investigations revealed that the lymphoid structure of the chicken Harderian gland is organized in different histological frameworks. In the head the surface epithelium of the central canal can be classified as a lymphoepithelial tissue which covers the dense lymphoid substance. It consists of small and medium-sized lymphocytes, dendritic-like cells, and occasional macrophages. High endothelial venules are associated with intense lymphocyte migration and homing that gives circumstantial evidence for a T-dependent region, as found in a secondary lymphoid organ. The B-dependent germinal centers are also common structural units of the head region's lymphoid substance. The body of the gland is loaded with plasma cells of different maturation stages. They immigrate into the epithelium of the central canal and produce IgM and IgA. Only a few scattered IgG producing plasma cells can be found in the gland of Harder. This plasmocytic region accounts for the immunosurveillance on the conjunctiva and in the upper respiratory tract through antibody production against bacterial or parasitic infections. In both the head and body regions of the gland, anti-B-L (anti-Ia) antibody recognized scattered elongated cells which might represent dendritic cells. The immunological relationship between the two histologically different parts of the Harderian gland is unknown, but we speculate that the dense lymphoid tissue with high endothelial venule receives the blood-borne, immunologically mature, but uncommitted B cells. By the influence of local antigen stimulus, these B cells transform to plasma cells which gradually appear in the body of the gland. The lymphoid structures of the head and the body fulfill the function of secondary and tertiary lymphoid organs, respectively.