Lymphocyte-mediated Responses Research Articles

Fast and slow releasing sulphide donors engender distinct transcriptomic alterations in Atlantic salmon hepatocytes

Hydrogen sulphide (H2S) is a naturally occurring compound generated either endogenously or exogenously and serves both as a gaseous signalling molecule and an environmental toxicant. Though it has been extensively investigated in mammalian systems, the biological function of H2S in teleost fish is poorly identified. Here we demonstrate how exogenous H2S regulates cellular and molecular processes in Atlantic salmon (Salmo salar) using a primary hepatocyte culture as a model. We employed two forms of sulphide donors: the fast-releasing salt form, sodium hydrosulphide (NaHS) and the slow-releasing organic analogue, morpholin-4-ium 4-methoxyphenyl(morpholino) phosphinodithioate (GYY4137). Hepatocytes were exposed to either a low (LD, 20 µg/L) or high (HD, 100 µg/L) dose of the sulphide donors for 24 hrs, and the expression of key sulphide detoxification and antioxidant defence genes were quantified by qPCR. The key sulphide detoxification genes sulfite oxidase 1 (soux) and the sulfide: quinone oxidoreductase 1 and 2 (sqor) paralogs in salmon showed pronounced expression in the liver and likewise responsive to the sulphide donors in the hepatocyte culture. These genes were ubiquitously expressed in different organs of salmon as well. HD-GYY4137 upregulated the expression of antioxidant defence genes, particularly glutathione peroxidase, glutathione reductase and catalase, in the hepatocyte culture. To explore the influence of exposure duration, hepatocytes were exposed to the sulphide donors (i.e., LD versus HD) either transient (1h) or prolonged (24h). Prolonged but not transient exposure significantly reduced hepatocyte viability, and the effects were not dependent on concentration or form. The proliferative potential of the hepatocytes was only affected by prolonged NaHS exposure, and the impact was not concentration dependent. Microarray analysis revealed that GYY4137 caused more substantial transcriptomic changes than NaHS. Moreover, transcriptomic alterations were more marked following prolonged exposure. Genes involved in mitochondrial metabolism were downregulated by the sulphide donors, primarily in NaHS-exposed cells. Both sulphide donors influenced the immune functions of hepatocytes: genes involved in lymphocyte-mediated response were affected by NaHS, whereas inflammatory response was targeted by GYY4137. In summary, the two sulphide donors impacted the cellular and molecular processes of teleost hepatocytes, offering new insights into the mechanisms underlying H2S interactions in fish.

Designing a novel SOX9 based multi-epitope vaccine to combat metastatic triple-negative breast cancer using immunoinformatics approach.

Immunotherapies are a promising treatment option especially for the management of TNBC owing to its higher levels of tumour-associated antigens together with higher mutational load. Of note, the administration of preventive vaccines in the early stage of the cancer holds promise for effective disease management. Therefore, the present study aimed to develop a novel multi-epitope peptide-based vaccination against TNBC employing SOX9, which has recently been recognized as a key regulator of TNBC metastasis. The immunodominant regions from the SOX9 protein were computed and assessed based on their ability to elicit both T and B lymphocyte mediated responses. The resultant epitopes were fused using appropriate linkers (EAAAK, KK, AAY and GPGPG) and adjuvant (50S ribosomal protein L7/L12) to enhance the vaccine’s immunogenicity. The physicochemical properties and population coverage were also anticipated for the constructed vaccine. Adding together, docking and dynamics simulation studies were performed on the modelled vaccine against TLR-4 to provide insight into the stability. Finally, the designed vaccine was cloned into the pET28 (+) vector and immunological simulation studies were carried out. These results demonstrate that our designed vaccine had the potency to trigger humoral and cellular immune responses. Based on these collective evidences, the final proposed vaccine could be an interesting therapeutics for the management of TNBC in the near future.Graphical abstractSchematic representation of an efficient vaccine design framework by combining the range of immunoinformatics strategies.Supplementary InformationThe online version contains supplementary material available at 10.1007/s11030-022-10539-w.

Immunoinformatics approach to designing a multi-epitope vaccine against Saint Louis Encephalitis Virus

The Saint Louis Encephalitis Virus (SLEV) is one of the causes of a rare, inflammatory condition of the brain tissues known as encephalitis. Belonging to the Flaviviridae family, SLEV can cause severe, detrimental repercussions on the central nervous system, leaving it impaired permanently. This study aimed to design and propose a multi-epitope vaccine candidate for preventing SLEV associated nervous system disorders. In this study, we used in silico approaches to predict potent epitopes on the envelope protein of SLEV by using multiple immunoinformatics and bioinformatics databases. We selected a total of 13 epitopes from the target envelope protein of SLEV through assessing their potential of eliciting both innate and acquired immunity by T and B lymphocyte mediated responses. Since SLEV is an RNA virus, conservancy of the epitopes were taken into account and the selected epitopes were found to be 100% conserved. The final multi-epitope vaccine subunit exhibited an antigenic score of 0.6797. Molecular docking of the multi-epitope vaccine construct was done with Toll-like receptor 4 (TLR4) protein and the energy score for the best model was found to be −1092.3. Expression capacity of the multi-epitope vaccine construct was tested in pET-28a (+) plasmid vector of Escherichia coli (strain-K12). Although the computational assays used in this study returned defensible results, further validation of the proposed vaccine candidate is required through in vitro and in vivo experiments to comment on its circumstantial efficacy. • Novel and potent chimeric vaccine against Saint Louis Encephalitis Virus . • Envelope proteins were selected by proteome exploration. • Immunoinformatics tools were used to identify potential T and B cell epitopes. • 99.97% worldwide population coverage was found for CTL and HTL epitopes. • Docking and dynamics simulation study revealed efficient binding and proper stability.

Upregulation of PD-L1 on Myeloma Cells By Immunomodulatory Agents Potentiates the Effect of Durvalumab

Introduction: Programmed death-ligand 1 (PD-L1) shows increased expression on plasma cells from multiple myeloma (MM) patients as compared with those from monoclonal gammopathy of undetermined significance patients and healthy volunteers. PD-L1 inhibits tumor-specific cytotoxic T lymphocyte-mediated responses via interaction with PD-1. Our previous study showed that PD-L1 on MM cells is associated with aggressive myeloma behaviors including cell proliferation and drug resistance, and furthermore the interaction of PD-L1 with PD-1 molecules induced drug resistance in MM cells by antiapoptotic responses through the Akt signaling pathway (Tamura et al. Leukemia 2013, Cancer Immunol Res 2016). We next investigated the regulation of PD-L1 expression by immunomodulatory drugs (IMDs) and antimyeloma effects of the anti-PD-L1 antibody durvalumab (DUR) in combination with IMDs.Methods: 1) The expression of mRNA and protein in MM cell lines (MOSTI-1, U266, KMS18, OPM2, MM.1S) and plasma cells from MM patients was analyzed using real-time PCR/DNA microarray and flow cytometry/Western blotting, respectively. 2) PD-L1 induction by IMDs was evaluated based on promoter activity in the luciferase assay. 3) Cereblon (CBRN)-knockdown and -knockout MM cell lines were established using the lentiviral shRNA and CRISPR/Cas9 system, respectively. 4) MM.1S cells were inoculated subcutaneously into immunodeficient NOD/Shi-scid, IL-2Rγnull (NOG) mice and then MM1.S-engrafted mice were administered lenalidomide (LEN; 25 mg/kg per day) orally for 12 days. 5) CD3+ T cells were isolated from peripheral blood mononuclear cells of healthy volunteers, and then T cells activated with anti-CD3/CD28 antibodies were co-cultured with X-irradiated MM cells for 4 days. T-cell proliferation and activation were assessed using the CFSE cell division assay and expression levels of activation markers CD25 and CD45RO, respectively.Results: 1) PD-L1 expression was upregulated in MM cell lines by LEN and pomalidomide (POM) in a concentration-dependent manner, but not by thalidomide. Its expression on plasma cells, but not on lymphocytes and monocytes, obtained from relapsed/refractory MM patients was enhanced by LEN and POM. This PD-L1 induction was not seen in CRBN-knockdown and -knockout MM cell lines. To clarify this mechanism, gene expression profiling analysis was performed in MM cell lines with and without POM treatment, demonstrating that not only PD-L1 but also a proliferation-inducing ligand (APRIL) was enhanced by POM. APRIL expression was not induced by POM in CRBN-knockdown and -knockout MM cell lines. PD-L1 upregulation on MM cells by POM was inhibited by either recombinant B cell maturation antigen (BCMA)-Ig as an APRIL inhibitor, the antibody against BCMA, one of the receptors for APRIL, or MEK/ERK inhibitor U0126. Furthermore, PD-L1 and APRIL expression in MM.1S cells engrafted in NOG mice were upregulated by LEN treatment. On the other hand, LEN/POM also increased PD-L1 promoter activity in BCMA- and APRIL-negative HeLa cells. 2) When T cells were co-cultured with LEN- or POM-treated U266 cells (PD-L1-upregulated cells), the proliferation of CD4+ and CD8+ T cells was inhibited in comparison with that in untreated cells. This T-cell suppression did not occur when DUR and LEN/POM were added during co-cultivation of LEN/POM-treated U266 cells with T cells. Moreover, the population of both CD25+CD45RO+ double-positive CD4+ and CD8+ T cells co-cultured with POM-treated U266 cells was increased by combined POM and DUR treatment compared with DUR alone.Conclusion: Our study revealed that IMDs enhanced PD-L1 expression on MM cells as well as APRIL production through CRBN binding, and one mechanism may involve the APRIL-BCMA interaction. Another mechanism may be directly involved in PD-L1 downregulation by IKZF1 degradation through CRBN. Furthermore, T-cell inhibition induced by PD-L1-upregulated MM cells was effectively recovered after combination treatment with DUR and LEN or POM. These results suggest that DUR combined with IMDs may be a reasonable treatment for relapsed/refractory MM patients. DisclosuresIto:Celgene: Honoraria. Handa:JVC KENWOOD Corporation: Other: Leadership Position/Advisory Role; SYSMEX CORPORATION: Other: Leadership Position/Advisory Role; TAMAGAWA SEIKI Co.,Ltd.: Other: Leadership Position/Advisory Role; Celgene: Honoraria, Other: Supplied with reagents and drugs., Research Funding. Tamura:Takeda Pharmaceutical: Honoraria; Ono Pharmaceutical: Honoraria; Bristol-Myers Squibb: Honoraria; Celgene: Honoraria, Research Funding.

Defining lymphocyte-predominant breast cancer by the proportion of lymphocyte-rich stroma and its significance in routine histopathological diagnosis.

Lymphocyte‐predominant breast cancer (LPBC) defined by the density of stromal lymphocytes shows favorable behavior. However, considerable distribution heterogeneity of lymphocytes is a major problem. The present study defined LPBC by the proportion of lymphocyte‐rich stroma with the cut‐off values of 30, 50, and 75%, and clinicopathologically analyzed mainly LPBC (area > 30%) defined by the cut‐off value of 30%. LPBCs (area > 30%), 39 cases in total, were composed mainly of triple‐negative and HER2+/ER ‐ subtypes, without any luminal A‐like subtype. LPBCs were composed predominantly of histological grade 3 tumors, without any grade 1 lesions. Multivariate analyses on 477 consecutive tumors revealed that ER‐negativity and grade 3 status associated significantly with LPBC. LPBC (area > 30%) showed better disease‐free survival than grade‐matched controls, and it was a good indicator of complete pathological remission after pre‐operative chemotherapy. Patients with LPBC with the cut‐off value of 50% and that of 75% showed 100% disease‐free survival. These results demonstrated the validity of our definition of LPBC. Our data also suggest that de‐differentiated cancers without TILs could be regarded as high‐grade cancer without lymphocyte‐mediated responses. In conclusion, the definition of LPBC by the proportion of lymphoid stroma is useful for prognostication of high grade breast cancer in routine diagnosis.

CORR Insights(®): do patients with a failed metal-on-metal hip implant with a pseudotumor present differences in their peripheral blood lymphocyte subpopulations?

T he study by Catelas and colleagues adds to the current scientific consensus that adaptive immune responses to metals occurs in a subset of all people with metal implants. The study authors also found that the incidence of pseudotumors correlates with systemic evidence of metal-related delayed-type hypersensitivity (DTH) responses and elevated metal ion levels. The current study identifies alterations in circulating adaptive immune cells in people with elevated levels of metal debris and pseudotumors. These findings support past histological and T-cell function studies, indicating pseudotumors are more prevalent in people with DTH-type immune reactivity. This highlights the growing recognition of these phenomena as central to implant performance in some individuals/implant designs. Case and group studies [4, 5] indicate that once locally activated, adaptive immune inflammation to metal contributes to (if not, mediates) the pathogenesis of poor implant performance. Metal DTH reactivity is a lymphocyte mediated response, similar to a Type IV DTH reaction. Metal-on-metal (MoM) hip failures have been associated with classic DTH responses including periimplant activated lymphocyte accumulations (aseptic lymphocytic vasculitisassociated lesions, [ALVAL]), and dermal and functional lymphocyte hypersensitivity responses to metals [2, 7, 11]. Elevated metal exposure can also cause different forms of toxicity responses in tissue, bone, and immune cells, including cobalt-induced hypoxia-like responses, postulated as causing pseudotumors [10]. Lymphocyte reactivity (metal DTH responses/ALVAL) and pseudotumors can be caused separately [6, 8]. Currently, there are no accepted thresholds for systemic metal ion levels that necessarily indicate for pseudotumor formation or immune reactions [1, 7]; it remains an increased risk association. This CORR Insights is a commentary on the article ‘‘Do Patients With a Failed Metal-onmetal Hip Implant With a Pseudotumor Present Differences in Their Peripheral Blood Lymphocyte Subpopulations?’’ by Catelas and colleagues available at: DOI: 10.1007/ s11999-015-4466-8. The author certifies that she, or any members of her immediate family, has no funding or commercial associations (consultancies, stock ownership, equity interest, patent/licensing arrangements, etc.) that might pose a conflict of interest in connection with the submitted article. All ICMJE Conflict of Interest Forms for authors and Clinical Orthopaedics and Related Research editors and board members are on file with the publication and can be viewed on request. The opinions expressed are those of the writers and do not reflect the opinion or policy of CORR or The Association of Bone and Joint Surgeons. This CORR Insights comment refers to the article available at DOI: 10.1007/s11999015-4466-8. N. J. Hallab PhD (&) Department of Orthopedic Surgery, Rush University Medical Center, 1735 W Harrison, Cohn Research Bldg, #722, Chicago, IL 60612, USA e-mail: nhallab@rush.edu CORR Insights Published online: 13 October 2015 The Association of Bone and Joint Surgeons1 2015

Iron Deficiency Impairs Intra-Hepatic Lymphocyte Mediated Immune Response.

Hepatic expression of iron homeostasis genes and serum iron parameters predict the success of immunosuppression withdrawal following clinical liver transplantation, a phenomenon known as spontaneous operational tolerance. In experimental animal models, spontaneous liver allograft tolerance is established through a process that requires intra-hepatic lymphocyte activation and deletion. Our aim was to determine if changes in systemic iron status regulate intra-hepatic lymphocyte responses. We used a murine model of lymphocyte-mediated acute liver inflammation induced by Concanavalin A (ConA) injection employing mice fed with an iron-deficient (IrDef) or an iron-balanced diet (IrRepl). While the mild iron deficiency induced by the IrDef diet did not significantly modify the steady state immune cell repertoire and systemic cytokine levels, it significantly dampened inflammatory liver damage after ConA challenge. These findings were associated with a marked decrease in T cell and NKT cell activation following ConA injection in IrDef mice. The decreased liver injury observed in IrDef mice was independent from changes in the gut microflora, and was replicated employing an iron specific chelator that did not modify intra-hepatic hepcidin secretion. Furthermore, low-dose iron chelation markedly impaired the activation of isolated T cells in vitro. All together, these results suggest that small changes in iron homeostasis can have a major effect in the regulation of intra-hepatic lymphocyte mediated responses.

Dormancy of growth-stunted malignant melanoma: sustainable and smoldering patterns.

The presentations of primary and metastatic cutaneous malignant melanoma (CMM) are very diverse. Evidence increasingly indicates that single CMM cells spread to distant sites quite early during cancer progression and are soon eliminated before they become clinically detectable. However bulky metastases which appear at a later stage might derive from some of these early neoplastic cells. It seems that local CMM single cell micro-metastases commonly predict sentinel lymph node involvement without overtly reflecting CMM progression to bulky visceral metastases. This study is intended to review the current understanding of the mechanisms underlying two CMM presentations. The first is the long interval, apparently disease-free, with persistent CMM dormancy, which may precede overt metastatic growth. Immunosurveillance may induce dormancy in single CMM cells disseminated in the body by blocking their proliferation cycle. The second is the so-called CMM smoldering phenomenon, which is marked by an alternate progression and regression of CMM locally with metastases that wax and wane for long periods of time over restricted skin areas. These very diverse patterns of CMM progression are likely to be ascribable to a number of biological factors, including the activation of CMM stem cells, and the combined phenotypic heterogeneity and variability in proliferative amplification in CMM cell clusters. Furthermore an adequate stimulation of CMM immune-surveillance and the induction of a specific stromal structure and vascular response are required. In this context, most early CMM tumors are in part controlled by lymphocyte-mediated responses before they become clinically detectable. However both the role of immune-surveillance and the mechanisms underlying both persistent and smoldering CMM dormancy remain unclear.

The Invasive Chytrid Fungus of Amphibians Paralyzes Lymphocyte Responses

The chytrid fungus, Batrachochytrium dendrobatidis, causes chytridiomycosis and is a major contributor to global amphibian declines. Although amphibians have robust immune defenses, clearance of this pathogen is impaired. Because inhibition of host immunity is a common survival strategy of pathogenic fungi, we hypothesized that B. dendrobatidis evades clearance by inhibiting immune functions. We found that B. dendrobatidis cells and supernatants impaired lymphocyte proliferation and induced apoptosis; however, fungal recognition and phagocytosis by macrophages and neutrophils was not impaired. Fungal inhibitory factors were resistant to heat, acid, and protease. Their production was absent in zoospores and reduced by nikkomycin Z, suggesting that they may be components of the cell wall. Evasion of host immunity may explain why this pathogen has devastated amphibian populations worldwide.

The Acute Phase of Chikungunya Virus Infection in Humans Is Associated With Strong Innate Immunity and T CD8 Cell Activation

Rapidly spreading to new regions, including the islands of the Indian Ocean, Central Africa, and Europe, Chikungunya fever is becoming a major problem of public health. Unlike other members of the alphavirus genus, immune responses to Chikungunya virus (CHIKV) have been poorly investigated. We conducted a large ex vivo multiplex study of 50 cytokine, chemokine, and growth factor plasma profiles in 69 acutely infected patients from the Gabonese outbreak of 2007. We also assessed a phenotypic study of T lymphocyte responses during human acute CHIKV infection. CHIKV infection in humans elicited strong innate immunity involving the production of numerous proinflammatory mediators. Interestingly, high levels of Interferon (IFN) α were consistently found. Production of interleukin (IL) 4, IL-10, and IFN-γ suggested the engagement of the adaptive immunity. This was confirmed by flow cytometry of circulating T lymphocytes that showed a CD8+ T lymphocyte response in the early stages of the disease, and a CD4+ T lymphocyte mediated response in the later stages. For the first time to our knowledge, we found evidence of CD95-mediated apoptosis of CD4+ T lymphocytes during the first 2 days after symptoms onset, ex vivo. Together, our findings suggest that strong innate immunity is required to control CHIKV infection.

Role of Dendritic Cells in the Induction of Proliferative T Lymphocyte-Mediated Responses to Iodine Contrast Media

Non-immediate allergic reactions to iodine contrast media (ICM) represent 2-5% of patients with adverse reactions to ICM. Although these reactions are T cell mediated, the lymphocyte transformation test (LTT) is frequently negative. We compared the LTT to ICM using either B lymphocytes and monocytes (B linf/mo: classical LTT) or Dendritic Cells (DC) as antigen presenting cells.

Foreign body response around total prosthetic metal-on-metal replacements of the temporomandibular joint in the UK

Replacements for the temporomandibular joint were developed in the early 1960s. Problems with various prostheses, notably the Kent VK1, led to detailed analysis of their risks and complications. In 1999 one type of prosthesis (the Christensen) was converted from an acrylic condyle on cobalt-chromium fossa to metal-on-metal cobalt-chrome condyle and fossa. This has been popular for the last 15 years in the UK, but since it was converted to the metal-on-metal variant there have been several cases of foreign body giant-cell reactions to the prosthesis. Of the 9 cases seen (out of 106 placed), 4 were found retrospectively to be sensitive to one of the metal components of the prosthesis; the others have not been tested to date. Other potential causes of this reaction are point contact, micromovement, or a lymphocyte-mediated response to the prosthesis.

LOOSENING AND OSTEOLYSIS ASSOCIATED WITH METAL-ON-METAL BEARINGS

LOOSENING AND OSTEOLYSIS ASSOCIATED WITH METAL-ON-METAL BEARINGS

The Association between Allergy and Rheumatoid Arthritis in the Canadian Population

Antagonism between Th1 and Th2 lymphocyte mediated responses has been proposed to explain the observed 20th Century population increase in Th2 mediated allergic disease and reciprocal decrease in infectious disease, which stimulates a Th1 mediated response. To determine if Th1/Th2 antagonism would be consistent with associations between non-infectious diseases, we tested the hypothesis that the population prevalence of Th2 mediated allergies is inversely related to the prevalence of Th1 mediated rheumatoid arthritis. The analysis was based on data from the Canadian Community Health Survey conducted by Statistics Canada in 2000-2001 of those at least 12 years of age from 125,129 households. Each subject was asked if he or she had certain chronic health conditions that had been diagnosed by a health professional. Logistic regression models were used to evaluate the association between rheumatoid arthritis and allergies with consideration of other important variables. Allergy history was positively related to the prevalence of rheumatoid arthritis both in women (adjusted odds ratio (OR): 1.57, 95% CI: 1.43, 1.73) and in men (adjusted OR: 1.55, 95% CI: 1.36, 1.77). The reported population prevalences of allergies and rheumatoid arthritis were positively associated and not explained by Th1/Th2 anatagonism suggesting that this mechanism may only be applicable to the association between an infectious and an immunologic disease. Mechanisms accounting for positive associations between immunologic diseases deserve further study.

Repeated exposure to silicone gel can induce delayed hypersensitivity.

The possible immunologic reactivity of silicone gel remains speculative and controversial. In this laboratory, a quantitative lymphocyte localization assay has been developed and well studied using pure lymphocytes collected by the technique of lymph vessel cannulation in sheep. The kinetics of antigen-specific immune responses (e.g., tuberculin reaction) in this model are well described and accepted. Using the known parameters regarding the response to purified protein derivative and the classic adjuvant Freund's Complete Adjuvant, this study was designed to identify the possible antigen-specific immunologic response, in the form of delayed-type hypersensitivity, after repeated exposure to silicone gel. Pure lymphocytes were collected by cannulating the efferent vessel of a subcutaneous lymph node in four groups of primed sheep which, 30 days previously, had received intradermal injections of 0.9% saline (negative controls; n = 6), Freund's Complete Adjuvant only (positive controls; n = 6), silicone gel (n = 7), or Freund's Complete Adjuvant homogenized with silicone gel (n = 7) in an attempt to induce sensitization. Multiple (1040) intradermal skin tests were performed using silicone gel, purified protein derivative, and 0.9% saline. After the skin lesions had developed for 48 hours, 5 x 10(8) lymphocytes were labeled in vitro with indium-111, returned intravenously, and allowed to circulate for 3 hours. Sheep were euthanized, the skin lesions were removed, and the radioactivity was counted in a gamma spectrometer. The radioactivity in each skin lesion is considered a measure of lymphocyte accumulation. The occurrence of augmented accumulation after reexposure to an antigen is a hallmark of delayed-type hypersensitivity. The purified protein derivative and saline lesions functioned as positive and negative controls, with counts per minute (cpm +/- standard error) of 2404 +/- 478 (Freund's Complete Adjuvant group) and 149 +/- 21 (saline group), respectively. Significantly greater (p = 0.0021) radioactivity was found in the silicone gel sites (310 +/- 35) in the silicone gel-primed group and the Freund's Complete Adjuvant plus silicone gel group (453 +/- 44; p = 0.0004) than in normal skin in each group. These data suggest that it may be possible to induce an antigen-specific lymphocyte-mediated response to silicone gel.

The Effect of In Vivo Interferon-Gamma on the Distribution of LFA-1 and ICAM-1 in Normal Human Skin

Lymphocyte function associated antigen 1 (LFA-1) and its ligand intercellular adhesion molecule 1 (ICAM-1) are cell surface adhesion molecules important in many lymphocyte-mediated responses. Recent in vitro studies have demonstrated that the cytokine interferon-gamma (IFN-gamma) can induce ICAM-1 expression by keratinocytes, and that lymphocytes adhere to IFN-gamma treated keratinocytes. In view of the importance of keratinocyte/lymphocyte interactions in the pathogenesis of cutaneous disease, we have examined the effects of in vivo IFN-gamma on cutaneous expression of LFA-1 and ICAM-1. Fourteen volunteers received intradermal IFN-gamma (dose: 1 or 10 micrograms) daily for 3 d. Biopsy was obtained on day 6. Cryostat sections were stained by the peroxidase antiperoxidase technique employing murine monoclonal antibodies to CD11, CD18, and ICAM-1. IFN-gamma intensified ICAM-1 expression by dermal endothelial cells and induced keratinocyte expression of ICAM-1. Furthermore, after administration of 10 micrograms of IFN-gamma LFA-1 positive (LFA + ve) lymphocytes were observed along the basement membrane zone closely related to ICAM-1 + ve basal keratinocytes and also surrounding dermal endothelium. Exposure to IFN-gamma induced expression of both CD11a and CD18 antigens on epidermal Langerhans cells. These studies suggest that the distribution of adherence molecules expression within cutaneous tissue in vivo is modulated by IFN-gamma, and that these alterations may be important in interactions involving cutaneous immunocompetent cells.

Evaluation of Antiviral Immunity Using Vaccinia Virus Recombinants Expressing Cloned Genes for Herpes Simplex Virus Type 1 Glycoproteins

Immunization of mice with vaccinia virus recombinants expressing the glycoproteins B or D of herpes simplex virus type 1 (HSV-1) induced humoral antibody as well as multiple aspects of HSV-1-specific T lymphocyte-mediated responses. However, vaccinated mice were not completely resistant to HSV-1 challenge and were unable to eliminate an epithelial infection rapidly. Evidence is presented which indicates that immunization with either vaccinia virus recombinant, while inducing the necessary protective populations of CD4+ T lymphocytes, fails to induce the complementing CD8+ cytotoxic T lymphocytes necessary for high levels of protection against a primary HSV-1 infection. These findings are discussed with relevance to the future development of anti-HSV vaccines.

Lymphocyte-mediated responses: activation of, and lysis by, cytotoxic lymphocytes

Lymphocyte-mediated responses: activation of, and lysis by, cytotoxic lymphocytes

Immune cell populations in cutaneous delayed-type hypersensitivity.

Delayed-type hypersensitivity (DTH) is a prototypic T lymphocyte-mediated response to antigenic challenge. In this study, mononuclear cells infiltrating the skin during cutaneous response to tuberculin in presensitized human subjects (responders) and nonimmune controls were identified using monoclonal antibodies by indirect immunofluorescence. In both responders and controls the infiltrate consisted mainly of T lymphocytes (T11+ and OKT3+) and monocytes (OKM1+, 63D3+, Mo2+) which initially accumulated in proximity to small blood vessels and later infiltrated the interstitial dermis and epidermis. More T lymphocytes reacted with OKT4 than with OKT8. 6 h after tuberculin the ratio of OKT4/OKT8 in tissue from responders exceeded that in blood, whereas in tissues studied at 15-48 h and in all control tissues those ratios in blood and tissue were similar. Evidence of T lymphocyte activation was sought using monoclonal antibodies anti-Tac, OKT9, and OKT10. In responders but not in controls the proportion of infiltrating cells reactive with these antibodies increased during the course of DTH. The presence of activated T lymphocytes in tissue was not associated with a comparable increase in peripheral blood cell populations identified by anti-Tac and OKT10. Studies using anti-B1, Leu-7, and anti-IgD/IgM revealed comparatively few reactive cells. Dual-labeling studies demonstrated that most Leu-7--reactive cells also bound T11 while fewer bound OKM1 or OKT8 and that cells reactive with OKIa1 and T11 constituted largely nonoverlapping populations. Specific patterns of reactivity were not observed when tissues were stained with anti-human C3, or poly C9-MA, a monoclonal antibody reactive with a neoantigen on polymerized C9 of the membrane attack complex of complement. The number of epidermal Langerhans cells identified by OKT6 was similar in responders and controls. Thus, the cutaneous response to tuberculin in sensitized individuals is characterized by early enrichment of the OKT4 subpopulation of T lymphocytes in tissue infiltrates and subsequent (15-48 h) evidence of T lymphocyte activation.

Antibody-dependent cellular protection against herpes simplex virus dissemination as revealed by viral plauqe and infectivity assays

Mouse nonimmune peripheral blood lymphocytes (PBL) plus antibody to herpes simplex virus inhibited virus dissemination in herpes simplex virus-infected 3T3 cell culture as revealed by development of viral plaque, size of immunofluorescent foci, appearance of polycaryocytes, and viral infectivity appearing in the culture. These nonimmune PBL did not act alone in inibiting virus dissemination, but did act synergistically in combination with antibody. The ratio of PBL to target monolayer cells needed to produce this effect was 20. Splenic lymphocytes had weak activity, whereas thymocytes were without effect, even in the presence of antibody. Neither interferon nor lymphotoxin was detected in this lymphocyte-mediated response. These findings support previous observations, based on cytotoxicity assays, that antibody-dependent cellular immune processes could be important in control of and recovery from herpesvirus infection.