Reduced Major Histocompatibility Complex Class Research Articles

IL-1β + TGF-β2 dual-licensed mesenchymal stem cells have reduced major histocompatibility class I expression and positively modulate tenocyte migration, metabolism, and gene expression.

The study objectives were to 1) determine the mesenchymal stem cell (MSC) surface expression of major histocompatibility complex (MHC) class I and transcriptome-wide gene expression changes following IL-1β + TGF-β2 dual licensing and 2) evaluate if IL-1β + TGF-β2 dual-licensed MSCs had a greater ability to positively modulate tenocyte function compared to naive MSCs. Equine bone marrow-derived MSCs from 6 donors and equine superficial digital flexor tenocytes from 3 donors. Experiments were performed in vitro. Flow cytometry and bulk RNA sequencing were utilized to determine naive and dual-licensed MSC phenotype and transcriptome-wide changes in gene expression. Conditioned media were generated from MSCs and utilized in tenocyte cell culture assays as a method to determine the effect of MSC paracrine factors on tenocyte function. Dual-licensed MSCs have a reduced expression of MHC class I and exhibit enrichment in functional pathways associated with the extracellular matrix, cell signaling, and tissue development. Additionally, dual-licensed MSC-conditioned media significantly improved in vitro tenocyte migration and metabolism to a greater degree than naive MSC-conditioned media. In tenocytes exposed to IL-1β, dual-licensed conditioned media also positively modulated tenocyte gene expression. Our data indicate that conditioned media containing paracrine factors secreted from dual-licensed MSCs significantly modulates in vitro tenocyte function, which may confer benefits in vivo to healing tendons following injury. Additionally, due to reduced MHC class I expression in dual-licensed MSCs, this technique may also provide an avenue to provide an effective "off-the-shelf" allogenic source of MSCs.

TRPA1 and TPRV1 Ion Channels Are Required for Contact Lens-Induced Corneal Parainflammation and Can Modulate Levels of Resident Corneal Immune Cells.

Contact lens wear can induce corneal parainflammation involving CD11c+ cell responses (24 hours), γδ T cell responses (24 hours and 6 days), and IL-17-dependent Ly6G+ cell responses (6 days). Topical antibiotics blocked these CD11c+ responses. Because corneal CD11c+ responses to bacteria require transient receptor potential (TRP) ion-channels (TRPA1/TRPV1), we determined if these channels mediate lens-induced corneal parainflammation. Wild-type mice were fitted with contact lenses for 24 hours or 6 days and compared to lens wearing TRPA1 (-/-) or TRPV1 (-/-) mice or resiniferatoxin (RTX)-treated mice. Contralateral eyes were not fitted with lenses. Corneas were examined for major histocompatibility complex (MHC) class II+, CD45+, γδ T, or TNF-α+ cell responses (24 hours) or Ly6G+ responses (6 days) by quantitative imaging. The quantitative PCR (qPCR) determined cytokine gene expression. Lens-induced increases in MHC class II+ cells after 24 hours were abrogated in TRPV1 (-/-) but not TRPA1 (-/-) mice. Increases in CD45+ cells were unaffected. Increases in γδ T cells after 24 hours of wear were abrogated in TRPA1 (-/-) and TRPV1 (-/-) mice, as were 6 day Ly6G+ cell responses. Contralateral corneas of TRPA1 (-/-) and TRPV1 (-/-) mice showed reduced MHC class II+ and γδ T cells at 24 hours. RTX inhibited lens-induced parainflammatory phenotypes (24 hours and 6 days), blocked lens-induced TNF-α and IL-18 gene expression, TNF-α+ cell infiltration (24 hours), and reduced baseline MHC class II+ cells. TRPA1 and TRPV1 mediate contact lens-induced corneal parainflammation after 24 hours and 6 days of wear and can modulate baseline levels of resident corneal immune cells.

Reduced MHC Class I and II Expression in HPV-Negative vs. HPV-Positive Cervical Cancers.

Cervical cancer (CC) is the second most common cancer in women worldwide and the fourth leading cause of cancer-associated death in women. Although human papillomavirus (HPV) infection is associated with nearly all CC, it has recently become clear that HPV-negative (HPV-) CC represents a distinct disease phenotype with increased mortality. HPV-positive (HPV+) and HPV- CC demonstrate different molecular pathology, prognosis, and response to treatment. Furthermore, CC caused by HPV α9 types (HPV16-like) often have better outcomes than those caused by HPV α7 types (HPV18-like). This study systematically and comprehensively compared the expression of genes involved in major histocompatibility complex (MHC) class I and II presentation within CC caused by HPV α9 types, HPV α7 types, and HPV- CC. We observed increased expression of MHC class I and II classical and non-classical genes in HPV+ CC and overall higher expression of genes involved in their antigen loading and presentation apparatus as well as transcriptional regulation. Increased expression of MHC I-related genes differs from previous studies using cell culture models. These findings identify crucial differences between antigen presentation within the tumor immune microenvironments of HPV+ and HPV- CC, as well as modest differences between HPV α9 and α7 CC. These differences may contribute to the altered patient outcomes and responses to immunotherapy observed between these distinct cancers.

Natural Killer Cells in Antibody Independent and Antibody Dependent HIV Control.

Infection with the human immunodeficiency virus (HIV), when left untreated, typically leads to disease progression towards acquired immunodeficiency syndrome. Some people living with HIV (PLWH) control their virus to levels below the limit of detection of standard viral load assays, without treatment. As such, they represent examples of a functional HIV cure. These individuals, called Elite Controllers (ECs), are rare, making up <1% of PLWH. Genome wide association studies mapped genes in the major histocompatibility complex (MHC) class I region as important in HIV control. ECs have potent virus specific CD8+ T cell responses often restricted by protective MHC class I antigens. Natural Killer (NK) cells are innate immune cells whose activation state depends on the integration of activating and inhibitory signals arising from cell surface receptors interacting with their ligands on neighboring cells. Inhibitory NK cell receptors also use a subset of MHC class I antigens as ligands. This interaction educates NK cells, priming them to respond to HIV infected cell with reduced MHC class I antigen expression levels. NK cells can also be activated through the crosslinking of the activating NK cell receptor, CD16, which binds the fragment crystallizable portion of immunoglobulin G. This mode of activation confers NK cells with specificity to HIV infected cells when the antigen binding portion of CD16 bound immunoglobulin G recognizes HIV Envelope on infected cells. Here, we review the role of NK cells in antibody independent and antibody dependent HIV control.

Reduced MHC class II expression in medullary thyroid cancer identifies patients with poor prognosis

Increasing body of recent studies determining the expression of tumor-specific major histocompatibility complex (MHC) class II protein supports its potential role in several malignancies, but little is known in human medullary thyroid cancer (MTC). Here, we report the expression of MHC-II and its clinicopathologic and prognostic relevance in MTC patients. Immunohistochemistry staining revealed a significant reduction in tumor cell-specific MHC-II expression in a higher AJCC stage and its poor prognostic correlation with human MTC development. Further statistical analysis identified the low MHC-II expression as a significant and independent risk factor for MTC recurrence and patient survival. Moreover, in vitro studies showed that the MHC-II expression was remarkably increased by RET inhibitors, which were prescribed to treat advanced MTC. Similarly, inhibitors blocking the MAPK/ERK and AKT/mTOR pathways also augmented MHC-II expression, suggesting their implications in RET-MHC-II signaling axis. Importantly, in vitro assays manifested enhanced peripheral blood leukocytes-mediated cytotoxicity in MTC cells treated with RET inhibitors, which were partially alleviated by HLA knock-down. Together, our study demonstrates that low MHC-II expression levels may serve as a prognostic biomarker for aggressive diseases in MTC patients and indicates that RET activation may promote MTC immune escape through downregulating MHC-II expression.

Reduced bonobo MHC class I diversity predicts a reduced viral peptide binding ability compared to chimpanzees

BackgroundThe highly polymorphic genes of the major histocompatibility complex (MHC) class I are involved in defense against viruses and other intracellular pathogens. Although several studies found reduced MHC class I diversity in bonobos in comparison to the closely related chimpanzee, it is unclear if this lower diversity also influences the functional ability of MHC class I molecules in bonobos. Here, we use a bioinformatic approach to analyze the viral peptide binding ability of all published bonobo MHC class I molecules (n = 58) in comparison to all published chimpanzee MHC class I molecules (n = 161) for the class I loci A, B, C and A-like.ResultsWe examined the peptide binding ability of all 219 different MHC class I molecules to 5,788,712 peptides derived from 1432 different primate viruses and analyzed the percentage of bound peptides and the overlap of the peptide binding repertoires of the two species. We conducted multiple levels of analysis on the “species”-, “population”- and “individual”-level to account for the characterization of MHC variation in a larger number of chimpanzees and their broader geographic distribution. We found a lower percentage of bound peptides in bonobos at the B locus in the “population”-level comparison and at the B and C loci in the “individual”-level comparison. Furthermore, we found evidence of a limited peptide binding repertoire in bonobos by tree-based visualization of functional clustering of MHC molecules, as well as an analysis of peptides bound by both species.ConclusionOur results suggest a reduced MHC class I viral peptide binding ability at the B and C loci in bonobos compared to chimpanzees. The effects of this finding on the immune defense against viruses in wild living bonobos are unclear. However, special caution is needed to prevent introduction and spread of new viruses to bonobos, as their defensive ability to cope with new viruses could be limited compared to chimpanzees.

MHC class I diversity in chimpanzees and bonobos

Major histocompatibility complex (MHC) class I genes are critically involved in the defense against intracellular pathogens. MHC diversity comparisons among samples of closely related taxa may reveal traces of past or ongoing selective processes. The bonobo and chimpanzee are the closest living evolutionary relatives of humans and last shared a common ancestor some 1 mya. However, little is known concerning MHC class I diversity in bonobos or in central chimpanzees, the most numerous and genetically diverse chimpanzee subspecies. Here, we used a long-read sequencing technology (PacBio) to sequence the classical MHC class I genes A, B, C, and A-like in 20 and 30 wild-born bonobos and chimpanzees, respectively, with a main focus on central chimpanzees to assess and compare diversity in those two species. We describe in total 21 and 42 novel coding region sequences for the two species, respectively. In addition, we found evidence for a reduced MHC class I diversity in bonobos as compared to central chimpanzees as well as to western chimpanzees and humans. The reduced bonobo MHC class I diversity may be the result of a selective process in their evolutionary past since their split from chimpanzees.

Molecular and cellular characteristics of human and non-human primate multipotent stromal cells from the amnion and bone marrow during long term culture.

IntroductionMultipotent stromal cells (MSCs) are among the key candidates in regenerative medicine. However variety of MSC sources and general heterogeneity lead to controversial data in functional characterization. Furthermore, despite intensive usage as preclinical animal model, little is known about MSCs of the common marmoset monkey.MethodsMSCs derived from placental amnion and bone marrow samples from human and common marmoset were characterized in parallel over 12 passages to monitor similarities and significant differences (p ≤ 0.05, Student’s t-test) in MSC markers and major histocompatibility complex (MHC) class I expression by immunohistochemistry, flow cytometry, real-time PCR, metabolic activity test, with special focus on pluripotency associated genes.ResultsHuman and non-human primate MSCs were characterized for expression of MSC markers and capability of differentiation into mesenchymal lineages. MSCs could be cultured more than 100 days (26 passages), but metabolic activity was significantly enhanced in amnion vs. bone marrow MSCs. Interestingly, MHC class I expression is significantly reduced in amnion MSCs until passage 6 in human and marmoset, but not in bone marrow cells. For MSC markers, CD73 and CD105 levels remain unchanged in amnion MSCs and slightly decline in bone marrow at late passages; CD166 is significantly higher expressed in human MSCs, CD106 significantly lower vs. marmoset. All cultured MSCs showed pluripotency marker expression like Oct-4A at passage 3 significantly decreasing over time (passages 6–12) while Nanog expression was highest in human bone marrow MSCs. Furthermore, human MSCs demonstrated the highest Sox2 levels vs. marmoset, whereas the marmoset exhibited significantly higher Lin28A values. Bisulfite sequencing of the Oct-4 promoter region displayed fewer methylations of CpG islands in the marmoset vs. human.ConclusionsLittle is known about MSC characteristics from the preclinical animal model common marmoset vs. human during long term culture. Studied human and common marmoset samples share many similar features such as most MSC markers and reduced MHC class I expression in amnion cells vs. bone marrow. Furthermore, pluripotency markers indicate in both species a subpopulation of MSCs with true ‘stemness’, which could explain their high proliferation capacity, though possessing differences between human and marmoset in Lin28A and Sox2 expression.

Endoplasmic reticulum aminopeptidase 1 and rheumatic disease: functional variation.

To review the recent developments in our understanding of endoplasmic reticulum (ER) aminopeptidase 1 (ERAP1) function in relation to its role in major histocompatibility complex (MHC) class I peptide presentation and human leukocyte antigen (HLA) class I-associated diseases. ERAP1 polymorphisms exhibiting loss-of-function have been associated with protection from AS. The aminopeptidase function of ERAP1 optimizes peptides for binding and presentation by MHC class I. Most of the studies have revealed reduced MHC class I expression in situations of reduced ERAP1 function. Under these circumstances, the presented peptides are often N-terminally extended, and cell surface complexes are unstable and fall apart more readily. In contrast, peptides presented by HLA-B*27 : 05 when ERAP1 is silenced are frequently extended on the C-terminus. Recent work has emphasized on the importance of assessing the function of allotypes encoded by ERAP1 haplotypes, rather than effects of single amino acid substitutions. The allotypes found in a series of AS patients were poorer at restoring HLA-B27 expression than allotypes found in unaffected controls, which may seem contrary to the genetic data linking loss-of-function to protection. More work is needed to understand how ERAP1 variants associated with risk and protection influence the quality and quantity of peptides available for binding to HLA class I molecules in the ER. Moreover, we need to determine allele-specific effects of ERAP1 variants in the context of HLA-B*51 and HLA-Cw*6, which are associated with Behçet's disease and psoriasis, respectively.

The Unfolded Protein Response (UPR)-activated Transcription Factor X-box-binding Protein 1 (XBP1) Induces MicroRNA-346 Expression That Targets the Human Antigen Peptide Transporter 1 (TAP1) mRNA and Governs Immune Regulatory Genes

To identify endoplasmic reticulum (ER) stress-induced microRNAs (miRNA) that govern ER protein influx during the adaptive phase of unfolded protein response, we performed miRNA microarray profiling and analysis in human airway epithelial cells following ER stress induction using proteasome inhibition or tunicamycin treatment. We identified miR-346 as the most significantly induced miRNA by both classic stressors. miR-346 is encoded within an intron of the glutamate receptor ionotropic delta-1 gene (GRID1), but its ER stress-associated expression is independent of GRID1. We demonstrated that the spliced X-box-binding protein-1 (sXBP1) is necessary and sufficient for ER stress-associated miR-346 induction, revealing a novel role for this unfolded protein response-activated transcription factor. In mRNA profiling arrays, we identified 21 mRNAs that were reduced by both ER stress and miR-346. The target genes of miR-346 regulate immune responses and include the major histocompatibility complex (MHC) class I gene products, interferon-induced genes, and the ER antigen peptide transporter 1 (TAP1). Although most of the repressed mRNAs appear to be indirect targets because they lack specific seeding sites for miR-346, we demonstrate that the human TAP1 mRNA is a direct target of miR-346. The human TAP1 mRNA 3'-UTR contains a 6-mer canonical seeding site for miR-346. Importantly, the ER stress-associated reduction in human TAP1 mRNA and protein levels could be reversed with an miR-346 antagomir. Because TAP function is necessary for proper MHC class I-associated antigen presentation, our results provide a novel mechanistic explanation for reduced MHC class I-associated antigen presentation that was observed during ER stress.

Identification of an alternate splice form of tapasin in human melanoma

Assembly of major histocompatibility complex (MHC) class I molecules with peptide in the endoplasmic reticulum requires the assistance of tapasin. Alternative splicing, which is known to regulate many genes, has been reported for tapasin only in the context of mutations. Here, we report on an alternate splice form of tapasin (tpsnΔEx3) derived from a human melanoma cell line that does not appear to be caused by mutations. Excision of exon 3 results in deletion of amino acids 70 to 156 within the beta barrel region, but the membrane proximal Ig domain, the transmembrane domain, and cytoplasmic tail of tapasin are intact. Introduction of tpsnΔEx3 into a tapasin-deficient cell line does not restore MHC class I expression at the cell surface. Similar to a previously described tapasin mutant (tpsnΔN50), tpsnΔEx3 interacts with TAP. Therefore, we used these altered forms of tapasin to test the importance of MHC class I interaction with TAP. In the presence of wild-type tapasin, transfection of tpsnΔN50, but not tpsnΔEx3, reduced MHC class I expression at the cell surface likely due its ability to compete MHC class I molecules from TAP. Together these findings suggest that tumor cells may contain alternate splice forms of tapasin which may regulate MHC class I antigen presentation.

Monocyte Major Histocompatibility Complex Class II Expression in Term and Preterm Labor

To investigate how term and preterm labor (PTL) influence the balance between maternal proinflammatory and antiinflammatory responses as measured by expression of major histocompatibility complex (MHC) Class II on maternal monocytes and tumor necrosis factor-alpha (TNF-alpha) production by in vitro stimulation of whole blood by lipopolysaccharide (LPS). Blood was taken from the following women (n=118): term elective cesarean delivery or in spontaneous labor, in premature labor, or with preterm premature rupture of the membranes (PROM) at less than 32 weeks, and gestation-matched reference group. Monocyte MHC Class II expression was measured by flow cytometry using a dual-staining technique. Plasma cytokine levels were assayed using a cytometric bead array system. In vitro whole blood stimulation with LPS was also performed, and cytokine production was measured. Term labor was associated with a fall in the percentage of monocytes expressing MHC Class II, compared with third trimester of pregnancy, P<.05 and a reduction in LPS-stimulated TNF-alpha production. This fall in MHC Class II was even more pronounced in PTL and preterm PROM groups compared with the reference group, P<.01. There was evidence of reduced expression of monocyte MHC Class II and LPS-stimulated TNF-alpha in term and preterm labor. This pattern of reduced MHC Class II expression and reduced TNF-alpha production is known as monocyte hyporesponsiveness or immune paresis. Detection of this state may provide insights into the maternal inflammatory status and be of use in the management of women with threatened PTL or preterm PROM. II.

High-risk human papillomavirus E7 expression reduces cell-surface MHC class I molecules and increases susceptibility to natural killer cells

High-risk human papillomavirus (HPV) is a major causative agent of cervical cancer and the E6 and E7 genes encode the major HPV oncoproteins. The E7 protein from high-risk HPV types alters cell cycle progression and represses genes encoding components of the antigen-presentation pathway, suggesting a role for E7 in tumour immune evasion. We show that knockdown of E7 expression in HPV16- and HPV18-transformed cervical carcinoma cells by RNA interference increased expression of major histocompatibility complex (MHC) class I at the cell surface and reduced susceptibility of these cells to natural killer (NK) cells. Tetracycline-regulated induction of HPV16 E7 resulted in reduced expression of cell surface MHC class I molecules and increased NK cell killing. Our results suggest that, for HPV-associated malignancies, reduced MHC class I expression is the result of an active immune evasion strategy that has evolved to assist viral replication.

The role of human leukocyte antigen E and G in HIV infection

The role of human leukocyte antigen E and G in HIV infection

Interferon regulatory factor-1 is required for interferon-γ-induced MHC class I genes in astrocytes

Recent studies have shown that the role of the transcription factor interferon regulatory factor-1 (IRF-1) in the expression of major histocompatibility complex (MHC) class I molecules is tissue-specific. Our previous studies indicated a role for IRF-1 in expression of MHC class I genes in cultured astrocytes in response to interferon-γ (IFN-γ). However, the requirement for IRF-1 in MHC class I expression has not been directly analyzed in neural tissue. To further ascertain the importance of IRF-1 in the induction of MHC class I genes in astrocytes in response to IFN-γ, we analyzed astrocytes from mice with a targeted disruption of the IRF-1 gene (IRF-1 −/− mice). As expected, astrocytes from wild type (IRF-1 +/+) mice showed a coordinate increase in both IRF-1 and MHC class I gene expression in response to IFN-γ. To the contrary, astrocytes from IRF-1 −/− mice had greatly reduced MHC class I mRNA expression. MHC class I gene promoter activity in astrocytes was controlled entirely through a single enhancer, the MHC-IRF-E, to which IRF-1 bound in response to IFN-γ in wild type but not in IRF-1 −/− mouse astrocytes. In vivo, astrocytes in brains of wild type mice readily responded to IFN-γ to express MHC class I molecules. This correlated with increased MHC class I mRNA in the brain. In contrast, brains of IRF-1 −/− mice showed no MHC class I gene induction following exposure to IFN-γ indicating that all cells in the central nervous system (CNS) including astrocytes with the potential to express MHC class I molecules were dependent on IRF-1. These studies conclusively demonstrate a major role for IRF-1/MHC-IRF-E interactions in controlling MHC class I gene expression in astrocytes in response to IFN-γ.

Impaired immune responses and altered peptide repertoire in tapasin-deficient mice.

Tapasin is a component of the major histocompatibility complex (MHC) class I antigen-loading complex. Here we show that mice with a disrupted tapasin gene display reduced MHC class I expression. Cytotoxic T cell (CTL) responses to viruses are impaired, and dendritic cells of tapasin-deficient mice do not cross-present protein antigen via the MHC class I pathway, indicating a defect in antigen processing. Natural killer (NK) cells from tapasin-deficient mice have an altered repertoire and are self-tolerant. In addition, the repertoire of class I-bound peptides is altered towards less stably binding ones. Thus tapasin plays a role in CTL and NK immune responses and in optimal peptide selection.

Trypanosoma brucei brucei infection impairs MHC class II antigen presentation capacity of macrophages.

During African trypanosomiasis, macrophages play a central role in T cell hyporesponsiveness to parasite-related and unrelated antigens. In this study, the ability of macrophages from Trypanosoma b. brucei-infected mice to present exogenous antigens to a major histocompatibility complex (MHC) class II-restricted CD4+ T cell hybridoma was analysed. We demonstrate that the antigen presentation capacity of macrophages from infected mice is markedly reduced as a result of a lower expression of [MHC class II-peptide] complexes on their plasma membrane. This defect did not result from a decreased antigen uptake/catabolism, a reduced MHC class II and intercellular adhesion molecule 1 expression on the surface of macrophages, a decreased affinity of MHC class II molecules for antigenic peptides, a competition between exogenous and parasite antigens, or the generation of inhibitory peptides. Our data indicate that the step resulting in coexpression of processed antigens and MHC class II molecules is affected in T. b. brucei-infected mice. Additionally, macrophages from infected mice secreted IL-10 that in turn contributes to the impairment of T cell activation.

Cytokine modulation of T-lymphocyte activation by intestinal smooth muscle cells

BACKGROUND & AIMS: Interleukin 1beta (IL-beta) and tumor necrosis factor alpha (TNF-alpha) are present in the neuromuscular layers during intestinal inflammation and directly affect intestinal smooth muscle function. We investigated whether IL-1beta and TNF-alpha modulate T- cell activation by murine intestinal smooth muscle cells (ISMCs). METHODS: alpha- and gamma- actin expression in ISMCs was confirmed using reverse-transcription polymerase chain reaction (RT-PCR). ISMCs were analyzed for class II major histocompatibility complex (MHC), intercellular adhesion molecule 1 (ICAM-1), and B7 before and after exposure to interferon gamma (IFN-gamma; 100 or 1000 U/ mL) in the presence or absence of IL-1beta (10 ng/mL) or TNF-alpha (5 ng/mL) for 72 hours. T-cell proliferation on cytokine-pretreated ISMCs was measured in the absence or presence of anti-B7 antibodies. RESULTS: In a dose-dependent fashion, IFN-gamma-pretreated ISMCs expressed MHC class II, ICAM-1, and B7-2, and stimulated T-cell proliferation. Pretreatment of ISMCs with IL-1beta and IFN-gamma reduced MHC class II and ICAM-1 expression and inhibited T-cell proliferation. When added with 100 U/mL IFN-gamma, TNF-alpha enhanced MHC class II and ICAM-1 expression on ISMCs and T-cell proliferation. However, TNF-alpha and 1000 U/mL IFN-gamma significantly decreased MHC class II expression and T-cell proliferation. Anti-B7-2 monoclonal antibody but not anti-B7-1 inhibited T-cell proliferative responses by >50%. CONCLUSIONS: Because IL-1beta, TNF-alpha, and T cells are present in the intestinal muscle layers during inflammation, these cytokines may serve to modulate the activation of T cells in this site. (Gastroenterology 1997 Jun;112(6):1986-95)

Herpes simplex virus protein targets for CD4 and CD8 lymphocyte cytotoxicity in cultured epidermal keratinocytes treated with interferon-gamma.

In early recurrent herpetic lesions, CD4 T lymphocytes are the predominant infiltrating cells, and keratinocytes expressing major histocompatibility complex (MHC) class II antigens, induced by interferon-gamma (IFN-gamma), are the major site of herpes simplex virus (HSV) replication. IFN-gamma pretreatment of human keratinocytes in vitro reduced MHC class I antigen down-regulation by HSV-1 infection and induced expression of HLA-DR that was unaltered by subsequent HSV-1 infection. Incubation of these infected keratinocytes with phosphonoacetic acid (PAA) almost completely inhibited expression of four major HSV glycoproteins, although expression of early proteins was not affected. Weak CD8 T lymphocyte cytotoxicity against IFN-gamma-stimulated, HLA-DR-expressing HSV-1-infected keratinocytes was consistently directed to the immediate early/early proteins (all 9 patients tested) but against late proteins to a lesser degree (4/9 patients). However, CD4 T lymphocyte cytotoxicity was much greater and directed predominantly against late HSV-1 glycoproteins (all 9 subjects tested) in these cells.

Characterization of the induction of persistence of major histocompatibility complex class II by hybrids of macrophages from Bacillus Calmette Guerin‐resistant mice

Peritoneal macrophages (M phi) from mice that are resistant to infection by Mycobacterium bovis (strain BCG) (Bcgr) can be induced to express major histocompatibility complex (MHC) class II glycoproteins (I-A) continuously upon treatment with 100 units of recombinant interferon-gamma (rIFN-gamma). In contrast, M phi from mice that are susceptible to BCG (Bcgs) express I-A transiently. Persistent expression of I-A does not require the continued synthesis of the glycoprotein. Thus, treatment with cycloheximide (CHX) reduces I-A expression by M phi that express I-A transiently but does not affect the expression of I-A that is persistently expressed. It was not possible, in these studies, to characterize the induction of persistence independent of MHC class II expression because of the 24-48 h required for MHC class II synthesis and cycling to the cell surface. During this time, persistence was also induced. To characterize persistence independent of MHC class II induction we have produced M phi-M phi somatic cell hybrids that express I-A constitutively by fusing cells from a Bcgs M phi cell line with M phi from Bcgr mice. Treatment of some of the hybrids with CHX reduced MHC class II expression. The M phi hybrids required treatment with high doses of rIFN-gamma to induce CHX-resistant I-A expression. The induction of the persistence of I-A, following the addition of rIFN-gamma, required a short burst of protein synthesis as well as the presence of rIFN-gamma for at least 3 h. The addition of actinomycin D simultaneously with rIFN-gamma did not prevent the induction of the persistence of I-A expression by one of the M phi hybrids (F6.4). In contrast, the induction of persistence of I-A expression required a longer period of induction than was observed for hybrid F6.4, which was attributed to the requirement for new RNA and protein synthesis by the A1.8 hybridoma.