Effects of T‐Lymphocyte‐Dependent and ‐Independent Immunity on Cholinergic Enzyme Activity in Mouse Lacrimal Gland

Abstract

The nature of the immune response following ocular immunization with a protein and a polysaccharide and the effects such immunization have on the activities of cholinergic enzymes in the lacrimal glands of BALB/c mice were examined. Lacrimal glands are highly innervated by sympathetic and parasympathetic nerve fibres and are involved in mucosal immunity and therefore are excellent sites to study neuro‐immune interactions. In this report, a T‐lymphocyte‐dependent protein antigen, keyhole limpet haemocyanin (KLH) and a T‐lymphocyte‐independent polysaccharide antigen, dextran (DEX) were administered topically to the eyes or intraperitoneally injected. Both routes of immunization produced a strong serum antibody response when KLH was the antigen. DEX, however, evoked a serum antibody response only after intraperitoneal administration. Eosin‐haematoxylin staining indicated no histological abnormality or inflammatory changes in any immunized lacrimal glands, but immuno‐staining revealed that only in the KLH‐treated tissues were IgG‐producing plasma cells discernible. Furthermore, KLH‐specific antibody was also detectable using an immuno‐blot assay in lacrimal glands. Polymerase chain reaction analysis with cytokine‐specific primers revealed induction of interleukin‐4 (lL‐4) in KLH‐treated lacrimal glands, but not in DEX or unimmunized tissues. Thus, the nature of the antigen seems important in the induction of the immune response in lacrimal glands. To delineate the effects that immunogenic differences might have on the activities of the cholinergic enzymes, choline acetyl‐transferase (ChAT) and acetylcholinesterase (AChE) were assayed using radiolabelled substrates and measuring labelled products. Both ChAT and AChE activities were influenced following KLH immunization, while DEX had only transient effects on ChAT. This is possibly due to the fact that KLH, a protein antigen, is the effective inducer of the specific immune response in the lacrimal gland, while DEX is not.

Autoreactive responses to an environmental factor: 1. phthalate induces antibodies exhibiting anti‐DNA specificity

“…Reagents and kits were from the following sources: Dulbecco's modified Eagle's minimal essential medium (DMEM), horse and fetal bovine serum (Gibco/Invitrogen, Grand Island, NY); Calf thymus DNA, oligonucleotide d(pT)4, rabbit anti‐mouse immunoglobulin–horseradish peroxidase (Ig‐HRP) reagent, o ‐phenylenediamine (OPD), methylated bovine serum albumin (BSA) and BSA (Sigma Chemical Co., St. Louis, MO); synthetic homopolymers, d(pA)10, d(pT)10, d(pG)10, and d(pC)10 (IDT, Santa Clara, CA); random hexamer primers, pd(N)6 (Amersham Pharmacia Biotech Inc. Piscataway, NJ); KLH (Calbiochem, CA); 4‐aminophthlate (Pfaltz and Bauer, Inc., Waterbury, CT); m ‐maleimidobenzoyl‐ N ‐hydroxysuccinimide ester (MBS), and Bradford protein assay kit (Pierce Chemicals, Rockford, IL); RNA isolation system (Biotecx Laboratory, Houston, TX), Maloney murine leukaemia virus reverse transcriptase (MMTV‐RT, from Epicenter Technologies, Madison, WI), Taq polymerase (Stratagene, La Jolla, CA). cDNAs from spleen of BALB/c and NZB/W F1 were obtained by RT–polymerase chain reaction (PCR) as previously described 18 . Other reagents used include an antioestriol mAb 1BF7 described before, 19 and normal mouse immunoglobulin M and 2C3‐Ig‐HRP prepared as described 20 .…”

“…cDNAs from spleen of BALB/c and NZB/W F1 were obtained by RT± polymerase chain reaction (PCR) as previously described. 18 Other reagents used include an antioestriol mAb 1BF7 described before, 19 and normal mouse immunoglobulin M and 2C3-Ig-HRP prepared as described. 20 2C3-Ig V H and V L complementarity determining regions and an unrelated control peptide CEMSDMSFSKDWSFYI were obtained from Alpha Diagnostic International, Inc. (San Antonio, TX).…”

Ocular Mucosal Immunity

“…Responses involving T lymphocyte-dependent antigens have been shown to result in production of IL-4 in lacrimal glands and influence cholinergic enzyme activity (Sinha et al, 2001), affecting immune processes and lacrimal physiology. Further, neuropeptides released into lymphoid structures or inflamed tissues are known to be chemotactic for APCs and to affect their interactions with T cells (Lambrecht, 2001).…”

Reconciling neuronally and nonneuronally derived acetylcholine in the regulation of immune function

“…However, no defect was observed when expansion of CD8 + T cells in M 1 or M 5 mAChR‐KO mice was stimulated by infection with lymphocytic choriomeningitis virus or vesicular stomatitis virus, which suggests that M 1 and M 5 mAChRs are not involved in antiviral immunity 60 . On the other hand, keyhole limpet hemocyanin, a protein antigen that induces both B cell– and T cell–mediated responses, upregulates the activities of both ChAT and AChE in mouse lacrimal gland, 61 suggesting that protein antigens do stimulate lymphocytic cholinergic activity. Furthermore, T cell activation induced by PHA, PMA (which activates PKC‐ and mitogen‐associated protein kinase pathways), or anti‐CD11a mAb upregulated both M 5 mAChR and ChAT gene expression (reviewed in Refs.…”