Lymphocyte Cell Numbers Research Articles

Abstract 589: Hedgehog signaling augments PDL-1 expression in cancer cells under hypoxic condition to inhibit antitumor effects by activated lymphocytes

Abstract [Background] Hedgehog (Hh) signaling is activated under hypoxic condition in cancerous tissue. This is thought to be one of the mechanisms of the induction of malignant phenotype of cancers. We have shown that Hh inhibitor decreases proliferation, invasiveness, and tumorigenesis. We have also revealed that Hh signaling plays pivotal roles for the maintenance of function in activated lymphocytes and dendritic cells under hypoxia. Recently, immune checkpoint inhibitor takes much attention, however, response rate is still limited. The mechanisms regulating PDL-1 expression also remains unclear. In this study, to increase reponse rate of immune checkpoint inhibitor, we investigated whether hypoxia-induced activation of Hh signaling contributes to PDL-1 expression in cancer and whether it affects the antitumor function of activated lymphocytes. [Materials and Methods] Pancreatic cancer cell line, Panc-1, gallbladder cancer cell line, NOZ and GBd, and lung small cell cancer cell line, SBC-5 were used as target cancer cells. Human lymphocytes derived from PBMC were activated by using anti CD3 mAb and IL-2, and were used as effector cells. For normoxic conditions, cells were cultured in 5% CO2 and 95% air. For hypoxic conditions, cells were cultured in 1% O2, 5% CO2, and 94% N2 using a multigas incubator. For Hh signaling inhibition, cyclopamine, polysaccharide-K (PSK), and small interfering RNA targeting Gli1, Smo, MAML3, and RBPJ were used. Cell numbers were counted by light microscope. Expression of cell surface molecules was estimated by FACS. [Results] 1) Hypoxia augmented PDL-1 expression in all 4 cancer cell lines. 2) Inhibition of Hh signaling using MAML3 siRNA, cyclopamine and PSK reduced PDL-1 expression under hypoxia in all 4 cancer cell lines. 3) When activated lymphocytes were cocultured with cancers treated with a Hh inhibitor, activated lymphocyte cell numbers significantly increased under hypoxia. 4) In contrast, this increase was abrogated when cancer cells were treated with a PDL-1 neutralizing antibody. 5) When activated lymphocytes were cocultured with cancers treated with a Hh inhibitor and/or anti-PDL-1 Ab, the percent of CD8+ lymphocytes decreased in both the cyclopamine- and anti-PDL-1-treated groups under hypoxic conditions, while there was no significant change in CD3 expression. 6) NKG2D expression increased on activated lymphocytes in anti PDL-1Ab-treated group. [Conclusion] These results suggest that Hh signaling is one of regulatory pathways of PDL-1 expression under hypoxia and that inhibiting Hh signaling may induce lymphocyte antitumor activity. Therefore, Hh inhibition could be a promising drug target, not only as a direct tumor suppressor agent, but also as a PDL-1 inhibitor. Citation Format: Hideya Onishi, Akiko Fujimura, Yasuhiro Oyama, Makoto Kawamoto, Akio Yamasaki, Takashi Morisaki. Hedgehog signaling augments PDL-1 expression in cancer cells under hypoxic condition to inhibit antitumor effects by activated lymphocytes [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 589. doi:10.1158/1538-7445.AM2017-589

The Effect of a combination of 12% spirulina and 20% chitosan on macrophage, PMN, and lymphocyte cell expressions in post extraction wound

Background: Tooth extraction is the ultimate treatment option for defective teeth followed by the need for dentures. Inflammation is one phase of the healing process that should be minimized in order to preserve alveolar bone for denture support. Macrophage, PMN and lymphocyte cells are indicators of acute inflammation. Spirulina and chitosan are natural compounds with the potential to be anti-inflammatory agents. Purpose: This research aimed to determine macrophage, PMN and lymphocyte cells of animal models treated with a combination of 12% spirulina and 20% chitosan on the 1st, 2nd and 3rd post-extraction day. Methods: Animal models were randomly divided into control (K) and treatment (P) groups. Each group was further divided into three subgroups (KI, KII, KIII and PI, PII, PIII). The post-extraction sockets of the control group animals were then filled with CMC Na 3%. Meanwhile, the post-extraction sockets of the treatment group members were filled with a combination of 12% spirulina and 20% chitosan. Subsequently, the number of PMN, macrophage and lymphocyte cells was analyzed by means of HE analysis on the 1st., 2nd. and 3rd. days. Statistical analysis was then performed using a T-test. Results: There was a decrease in PMN cells and an increase in macrophage and lymphocyte cells on Days 1, 2, and 3. Conclusion: It can be concluded that a combination of 12% spirulina and 20% chitosan can not only decrease PMN cells, but can also increase macrophage and lymphocyte cells on Days 1, 2 and 3 after tooth extraction.

Reactivity changes in the rats with streptozotocin-induced diabetes during chronic enterocolitis development

The results of the integral indices in diagnosis of endogenous intoxication in rats with diabetes and chronic enterocolitis with diabetes are presented. Streptozotocin-induced diabetes and chronic enterocolitis together with diabetes in rats are involved a violation of immunological reactivity. The use of carrageenan on the background of streptozotocin-induced diabetes in rats is accompanied by a left shift of leukocyte formula with the increasing of band neutrophils number and appearance of immature lymphocytic cells, the lowest segmented neutrophils and lymphocytosis.

Beauveria attenuates asthma by inhibiting inflammatory response and inducing lymphocytic cell apoptosis.

The present study aimed to investigate the role of beauveria (BEA) in asthma. We investigated the cytotoxic effect of BEA on the proliferation of inflammatory cells and secretion of inflammatory mediators both in-vitro and in-vivo. In in-vitro studies, BEA inhibited lymphocytic cell proliferation and the proliferation of lymphocytic cells induced by Phorbol-12-myristate-13-acetate (PMA). We used ELISA to test the effects of BEA on the secretion of inflammatory factors including tumor necrosis factor-alpha (TNF-α), interleukin-12 (IL-12) and interferon-gamma (IFN-γ). Flow cytometry was used to evaluate the influence of BEA on cell apoptosis. The effect of BEA on the cell numbers of eosinophils, lymphocytes, macrophages, neutrophils and other cells in mouse bronchoalveolar lavage fluid (BALF) was also evaluated. The expression of apoptosis related molecules Bax, Caspase-3 and Bcl-2 was examined by Western blotting analysis. Our results indicated that BEA played a protective role in asthma. BEA inhibited lymphocytic cell proliferation and secretion of inflammatory mediators. BEA promoted cell apoptosis, stimulated the expression of Bax and Caspase-3 and inhibited Bcl-2 protein expression in a dose-dependent manner. In in-vivo experiments, BEA reduced the cell number of eosinophils, lymphocytes, macrophages, neutrophils and other cells in mouse BALF. BEA inhibited secretion of inflammatory mediators, stimulated expression of Bax and Caspase-3, and inhibited expression of Bcl-2 in mouse lung tissue dose-dependently. All together, our results indicated that BEA could attenuate asthma by inhibiting inflammatory response and induce apoptosis of inflammatory cells.

Hedgehog signaling regulates PDL-1 expression in cancer cells to induce anti-tumor activity by activated lymphocytes

We investigated whether hypoxia-induced activation of Hh signaling contributes to PDL-1 expression in cancer and whether it affects the anti-tumor function of activated lymphocytes. Hypoxia augmented PDL-1 expression and inhibition of Hh signaling reduced PDL-1 expression under hypoxia. When activated lymphocytes were cocultured with cancers treated with a Hh inhibitor, activated lymphocyte cell numbers increased under hypoxia. In contrast, this increase was abrogated when cancer cells were treated with a PDL-1 neutralizing antibody. These results suggest that Hh signaling is one of regulatory pathways of PDL-1 expression under hypoxia and that inhibiting Hh signaling may induce lymphocyte anti-tumor activity.

MAP4K4 deficiency in CD4(+) T cells aggravates lung damage induced by ozone-oxidized black carbon particles.

As the main composition of combustion, black carbon (BC) is becoming more and more noticeable at home and abroad. Ozone-oxidized black carbon (oBC) was produced through aging of ozone, one of the near-surface pollutants, to black carbon. And oBC was found to be more oxidation and cell toxicity when compared with BC. Besides, as a key cell of immunity, whether CD4(+) T cell would involve in lung inflammation induced by particular matter is still unclear. This study aims to observe the effect of oBC on lung damage in mice and discuss how the functional MAP4K4 defect CD4(+) T cells (conditional knockout of MAP4K4) presents its role in this process. In our study, MAP4K4 deletion in CD4(+) T cells (MAP4K4 cKO) could increase cell number of macrophages, lymphocytes and neutrophils in bronchoalveolar lavage fluid (BALF) exposed to oBC. MAP4K4 deletion in CD4(+) T cell also affected CD4(+) T cell differentiation in mediastinal lymph nodes after oBC stimulation. The number of CD4(+) IL17(+) T cell increased obviously. The levels of IL-6 mRNA of lung in MAP4K4 cKO mice was higher than that in wild type mice after exposed to oBC, while the level of IL-6 in BALF had the same trend. Histological examination showed that MAP4K4 deletion in CD4(+) T cells affected lung inflammation induced by oBC. Results indicated that MAP4K4 cKO in CD4(+) T cells upgraded the level of inflammation in lung when exposed to oBC, which may be connected to the CD4(+) T cell differentiation and JNK, ERK and P38 pathways.

Elevated levels of FN1 and CCL2 in bronchoalveolar lavage fluid from sarcoidosis patients.

BackgroundSarcoidosis is a granulomatous systemic inflammatory disease in which more than 90 % of all patients develop pulmonary manifestations. Several gene associations have previously been described, but established and clinically useful biomarkers are still absent. This study aimed to find proteins in bronchoalveolar lavage (BAL) fluid that can be associated with the disease.MethodsWe developed and performed profiling of 94 selected proteins in BAL fluid and serum samples obtained from newly diagnosed and non-treated patients with sarcoidosis. Using multiplexed immunoassays, a total of 317 BAL and 217 serum samples were analyzed, including asthmatic patients and healthy individuals as controls.ResultsOur analyses revealed increased levels of eight proteins in sarcoidosis patients compared to controls. Out of these, fibronectin (FN1) and C-C motif chemokine 2 (CCL2) revealed the strongest associations. In addition, cadherin 5 (CDH5) was found to correlate positively with lymphocyte cell numbers in BAL fluid.ConclusionsApplying a high throughput proteomics screening technique, we found proteins of potential clinical relevance in the context of sarcoidosis.Electronic supplementary materialThe online version of this article (doi:10.1186/s12931-016-0381-0) contains supplementary material, which is available to authorized users.

Influence of transfusion of lymphokine-activated T killer cells on inflammatory responses in dogs after laparotomy.

The influence of transfusion of lymphokine-activated T killer cells (T-LAK) on inflammatory responses was examined in dogs after laparotomy. Plasma C-reactive protein (CRP) level, cell numbers of peripheral blood lymphocytes (PBLs) and T lymphocyte subsets (CD3+, CD4+ and CD8+) and mRNA expression levels of cytokines including interleukin (IL)-2, IL-12, IL-4, IL-10 and transforming growth factor (TGF)-β in peripheral blood mononuclear cells (PBMCs) were measured in dogs with (T-LAK group) or without (control group) a single T-LAK administration immediately after laparotomy. The plasma CRP level initially increased and then decreased to the normal range at 7 days after laparotomy in the T-LAK group, which was earlier than in the control group. The expression level of IL-10 mRNA showed a marked postoperative increase and was significantly higher than the preoperative level on day 7 (P<0.05), whereas the level in the control group showed no clear change after laparotomy. A significant increase in IL-2 mRNA expression level in the T-LAK group was observed on day 14, which was two weeks earlier than in the control group (P<0.05). These results suggest that T-LAK therapy in dogs after laparotomy leads to earlier resolution of postoperative inflammation by production of an anti-inflammatory cytokine (IL-10) in the early phase of the postoperative period and earlier restoration of cell-mediated immunity related to cytokine production by PBMCs.

Characterisation of haematological profiles and whole blood relative gene expression levels in Holstein-Friesian and Jersey bull calves undergoing gradual weaning

Haematological profiles indicate the health status of an animal and can be used to identify sub-clinical stress responses. The objectives of the study were to examine (i) the effect of breed and plane of nutrition, on haematological profiles of artificially reared Holstein-Friesian and Jersey bull calves in response to gradual weaning, and (ii) the effect of breed on immune response genes in bovine whole blood using real-time quantitative PCR. Holstein-Friesian and Jersey bull calves were group housed indoors and individually fed using an automatic feeder. They were allocated to a high, medium or low plane of nutrition, based on milk replacer (MR) and concentrate. The nutrition treatments were calculated using National Research Council guidelines in order to achieve a high, medium or low growth rate for each respective breed. During the weaning phase MR was gradually reduced over a 14-day (d) period (d −13 to d 0). Calves were blood sampled on d −14, −6, −3, 0, 1, 3, 8 and 14 relative to weaning (d 0) for subsequent haematological analysis. On d −14, 1 and 8, a subset of eight Holstein-Friesian calves randomly selected from the medium nutrition treatment and eight Jersey calves randomly selected from the high nutrition treatment, were blood sampled for gene expression profiling, targeting biomarkers of weaning stress. These two treatment groups were chosen to examine the effect of breed on expression of the genes of interest, as energy intake and animal performance were similar. There was no effect of breed×plane of nutrition interaction nor effect of plane of nutrition on any variable measured (P>0.05). Gradual weaning produced differential biological responses in the two breeds evidenced by breed×time interactions for lymphocyte, monocyte and red blood cell number, plasma haemoglobin and haptoglobin concentrations (P<0.05). The typical stress response consisting of neutrophilia and lymphopaenia was not observed for any treatment. An immune response to gradual weaning was observed as the relative gene expression level of the pro-apoptotic gene, Fas, increased on d 1 relative to d −14 (P<0.05). Relative gene expression levels were greater in Jersey calves compared with Holstein-Friesian for the pro-inflammatory cytokine CXCL8 (P=0.05) and the glucocorticoid receptor, GRα (P<0.05). The increased levels of these transcripts suggest that Jersey calves may have a more sensitive immune system compared with Holstein-Friesian.

Effect of floor type on hoof lesions, dirt scores, immune response and production of beef bulls

The objective of the study was to investigate the effect of old and new concrete slats (CS) with or without rubber mats (RM) on animal performance, hoof health, dirt scores and health status of finishing bulls. Continental crossbred beef bulls (n=72; mean initial live weight=441 (s.d. 45.1) kg) were blocked by breed and live weight and randomly assigned by block to one of four treatments; (1) Old CS (2) New CS, (3) Old CS covered with RM, (4) New CS covered with RM. Each treatment had 3 pens of 6 bulls at a space allowance of 2.9m2 per animal. Bulls were fed a total mixed ration (TMR) of silage and rolled barley on a 54:46 dry matter (DM) basis for 148 days. Feed was weighed into each pen daily and refusals were weighed twice weekly. Bulls were weighed every three weeks to coincide with dirt scoring. Total leucocyte, neutrophil, lymphocyte, eosinophil, monocyte, basophil and red blood cell number and haemoglobin concentrations were measured on day 0 and day 148. Bull's hooves were inspected for the presence of lesions at the start of the study and again at slaughter. After slaughter, carcass weight, carcass gain, conformation and fat score, kidney and channel fat and hide weight were recorded. Bulls on RM had a greater average daily gain (ADG) (0.16kg/day) (P<0.01), kidney and channel fat weight (KCF) (P<0.05), hide weight (P<0.01) and a better feed conversion ratio (FCR) (P<0.05) than those on CS. Floor type had no effect on dry matter intake (DMI), slaughter weight, carcass weight, kill out percentage, conformation score or fat score. Bulls on RM had 44% more hoof lesions (P<0.01) than those on CS. There were slat×time (P<0.05) and mat×time (P<0.01) interactions for dirt scores. Bulls on slats were dirtier than those on mats on days 63, 84 and 126 (P<0.01) while bulls on new slats were dirtier than those on old slats on days 21 (P<0.01) and 42 (P<0.05). Floor type had no effect (P>0.05) on any of the haematological variables measured which suggests that the immunological status of the bulls was not affected by treatment. While there was no evidence of lameness in bulls on RM, the increased number of hoof lesions suggests that hoof health may be compromised in bulls housed on RM.

Growth performance, serum biochemical profile, jejunal morphology, and the expression of nutrients transporter genes in deoxynivalenol (DON)- challenged growing pigs.

BackgroundFusarium infection with concurrent production of deoxynivalenol (DON) causes an increasing safety concern with feed worldwide. This study was conducted to determine the effects of varying levels of DON in diets on growth performance, serum biochemical profile, jejunal morphology, and the differential expression of nutrients transporter genes in growing pigs.ResultsA total of twenty-four 60-day-old healthy growing pigs (initial body weight = 16.3 ± 1.5 kg SE) were individually housed and randomly assigned to receive one of four diets containing 0, 3, 6 or 12 mg DON/kg feed for 21 days. Differences were observed between control and the 12 mg/kg DON treatment group with regards to average daily gain (ADG), although the value for average daily feed intake (ADFI) in the 3 mg/kg DON treatment group was slightly higher than that in control (P<0.01). The relative liver weight in the 12 mg/kg DON treatment group was significantly greater than that in the control (P<0.01), but there were no significant differences in other organs. With regard to serum biochemistry, the values of blood urea nitrogen (BUN), alkaline phosphatase (ALP), alanine aminotransferase (ALT) and aspartate amino transferase (AST) in the 3 treatment groups were higher than those in the control, and the serum concentrations of L-valine, glycine, L-serine, and L-glutamine were significantly reduced in the 3 treatment groups, especially in the 12 mg/kg DON group (P<0.01). Serum total superoxide dismutase (T-SOD), glutathione peroxidase (GSH-Px) were markedly decreased after exposure to DON contaminated feeds (P<0.01). The villi height was markedly decreased and the lymphocyte cell numbers markedly increased in the 3 DON contaminated feeds (P<0.01). The mRNA expression levels of excitatory amino acid transporter-3 (EAAC-3), sodium-glucose transporter-1 (SGLT-1), dipeptide transporter-1 (PepT-1), cationic amino acid transporter-1 (CAT-1) and y+L-type amino acid transporter-1 (LAT-1) in control were slightly or markedly higher than those in the 3 DON treatment groups.ConclusionsThese results showed that feeds containing DON cause a wide range of effects in a dose-dependent manner. Such effects includes weight loss, live injury and oxidation stress, and malabsorption of nutrients as a result of selective regulation of nutrient transporter genes such as EAAC-3, SGLT-1, PepT-1, CAT-1 and LAT-1.

Abstract B10: LFA-1/ICAM-1 interaction switches on an orchestrated prometastatic microenvironmental shift during experimental liver metastasis of colon C26 cancer cells.

Abstract Intercellular adhesion molecule (ICAM)-1 is the main counter-receptor for the lymphocyte function associated antigen (LFA)-1. However, the role this interaction plays during liver metastasis of colorectal cancer cells and its underlying mechanisms remain unclear. Previously, we showed that mannose receptor (ManR) stimulation on liver sinusoidal endothelial cells (LSECs) resulted in a reduced cytotoxic potential of liver infiltrating lymphocytes (LIL) towards C26 murine colon carcinoma cells. Using ICAM-1 specific siRNAs in vivo, we aimed to study the role of hepatic ICAM-1 in the prometastatic shift of the liver microenvironment. Livers of control and silenced mice, 24 hours and 14 days after C26 cell inoculation, were compared for: percentage of detained C26 cells, number and volume of metastatic foci, levels of recruited immune, myofibroblastic and inflammatory cells and whole liver expression of pro-inflammatory genes. Detained cancer cells and metastatic foci number were reduced in silenced mice compared to controls, along with a decrease in myeloid suppressor cells and recruited macrophages and myofibroblasts. On the contrary, significant higher numbers of lymphocytic cells were observed in mice with reduced ICAM-1. Finally, silencing of ICAM1 resulted in altered total liver mRNA levels of proinflammatory factors in the tumor-bearing mice. Therefore, an orchestrated prometastatic program might be switched on in the liver microenvironment by LFA-1/ICAM-1 interaction which, in turn, could lead not only to a reduced antitumor citotoxicity but, also, to activation of the stromal compartment favoring tumor progression, and, opening liver's doors to tumor cells to aggressively metastasize. Based on these results, LFA-1/ICAM-1 interaction arises as a potential therapeutic target in cancer treatment. Citation Format: Aitor Benedicto, Joana Marquez, Elvira Olaso, Beatriz Arteta. LFA-1/ICAM-1 interaction switches on an orchestrated prometastatic microenvironmental shift during experimental liver metastasis of colon C26 cancer cells.. [abstract]. In: Abstracts: AACR Special Conference on Cellular Heterogeneity in the Tumor Microenvironment; 2014 Feb 26-Mar 1; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2015;75(1 Suppl):Abstract nr B10. doi:10.1158/1538-7445.CHTME14-B10

Pro-inflammatory potential ofEscherichia colistrains K12 and Nissle 1917 in a murine model of acute ileitis

Non-pathogenic Escherichia coli (Ec) strains K12 (EcK12) and Nissle 1917 (EcN) are used for gene technology and probiotic treatment of intestinal inflammation, respectively. We investigated intestinal colonization and potential pro-inflammatory properties of EcK12, EcN, and commensal E. coli (EcCo) strains in Toxoplasma (T.) gondii-induced acute ileitis. Whereas gnotobiotic animals generated by quintuple antibiotic treatment were protected from ileitis, mice replenished with conventional microbiota suffered from small intestinal necrosis 7 days post-T. gondii infection (p.i.). Irrespective of the Ec strain, recolonized mice revealed mild to moderate histopathological changes in their ileal mucosa. Upon stable recolonization with EcK12, EcN, or EcCo, development of inflammation was accompanied by pro-inflammatory responses at day 7 p.i., including increased ileal T lymphocyte and apoptotic cell numbers compared to T. gondii-infected gnotobiotic controls. Strikingly, either Ec strain was capable to translocate to extra-intestinal locations, such as MLN, spleen, and liver. Taken together, Ec strains used in gene technology and probiotic treatment are able to exert inflammatory responses in a murine model of small intestinal inflammation. In conclusion, the therapeutic use of Ec strains in patients with broad-spectrum antibiotic treatment and/or intestinal inflammation should be considered with caution.

Investigation of immunological changes in mice subjected to animal model of sepsis

Sepsis induces multiple organ dysfunction syndrome including septic encephalopathy, that is commonly seen in systemically ill patients. Moreover, knowing that changes in the nervous system are implicated in the induction or amplification of immune system changes, or conversely, and they are not well elucidated in sepsis, we investigate the changes in brain function and immune system of mice in an animal model of sepsis by cecal ligation and puncture (CLP). Balb/c mice underwent CLP and were divided into two groups: CLP or Sham operation. We evaluated brain changes by SHIRPA behavioral test, after 6h of CLP. Mice were sacrificed, spleen were isolated and evaluated by flow cytometry profile of population of lymphocytes. We observe that the neuropsychiatric state in SHIRPA test was significantly decreased in animals with CLP. Regarding lymphocytes, no difference in the number of cells (B lymphocytes, regulatory T cells, helper T lymphocytes and cytotoxic T lymphocytes) compared to Sham and CLP. Thus, we conclude from these preliminary results that the decrease in neuropsychiatric condition is not correlated with cell number of lymphocytes. Support: FAPESP 2012/07898–0

Intravenous immunoglobulin treatment recovers the down-regulated levels of Th1 cytokines in the sera and skin of scleroderma patients

Systemic sclerosis (SSc) is an autoimmune disease characterized by fibrosis of the skin and internal organs. There is an urgent need to develop new therapeutic approaches against skin fibrosis. Although intravenous immunoglobulin (IVIG) may be one of the promising treatments, the mechanisms by which IVIG improves the fibrosis of SSc remain unknown. To compare the cytokine profile in the sera and skin of SSc patients before and after IVIG administration, and try to clarify the mechanism of the effect of IVIG. Cytokine array revealed that the serum levels of IFN-γ and IL-12, representative Th1 cytokines, were increased by IVIG treatment, but not by placebo. The percentage of IFN-γ- and IL-12-positive cell number/CD4-positive T lymphocyte cell number was also significantly increased by IVIG in SSc skin. Furthermore, mRNA expression of IFN-γ and IL-12 in SSc skin tissue was significantly up-regulated after IVIG treatment. The expression of Th1 cytokine is reported to be decreased in SSc. Our study suggested IVIG recovered the suppressed levels of Th1 cytokines, and that the treatment improves skin fibrosis by correcting the Th1/Th2 balance. In order to facilitate the clinical use of IVIG for SSc, it is necessary to perform a lager study in the future. JSID AbstractsJournal of Dermatological ScienceVol. 69Issue 2Preview Full-Text PDF

Behavioral, Virologic, and Immunologic Factors Associated With Acquisition and Severity of Primary Epstein–Barr Virus Infection in University Students

University students were studied prospectively to determine the incidence of and risk factors for acquisition of primary Epstein-Barr virus (EBV) infection and the virologic and immune correlates of disease severity. EBV antibody-negative freshmen participated in monthly surveillance until graduation. If antibodies developed, proximate samples were assayed for viral load by polymerase chain reaction. Lymphocyte and natural killer (NK) cell numbers and activation were measured by flow cytometry, and plasma cytokine levels were measured by a multiplex assay. Of 546 students screened, 202 (37%) were antibody negative; 143 antibody-negative students were enrolled. During a median of 3 years of observation, 66 subjects experienced primary infection. Of these, 77% had infectious mononucleosis, 12% had atypical symptoms, and 11% were asymptomatic. Subjects reporting deep kissing with or without coitus had the same higher risk of infection than those reporting no kissing (P < .01). Viremia was transient, but median oral shedding was 175 days. Increases were observed in numbers of NK cells and CD8(+) T-cells but not in numbers of CD4(+) T-cells during acute infection. Severity of illness correlated positively with both blood EBV load (P = .015) and CD8(+) lymphocytosis (P = .0003). Kissing was a significant risk for primary EBV infection. A total of 89% of infections were symptomatic, and blood viral load and CD8(+) lymphocytosis correlated with disease severity.

Intravenous immunoglobulin treatment recovers the down-regulated levels of Th1 cytokines in the sera and skin of scleroderma patients

BackgroundSystemic sclerosis (SSc) is an autoimmune disease characterized by fibrosis of the skin and internal organs. There is an urgent need to develop new therapeutic approaches against skin fibrosis. Although intravenous immunoglobulin (IVIG) may be one of the promising treatments, the mechanisms by which IVIG improves the fibrosis of SSc remain unknown.ObjectivesTo compare the cytokine profile in the sera and skin of SSc patients before and after IVIG administration, and try to clarify the mechanism of the effect of IVIG.MethodsEach three patients received 5-day administration of IVIG, or the same dose of physiologic saline for placebo. Cytokine levels were determined by ELISA array, immunostaining, and real-time PCR.ResultsCytokine array revealed that the serum levels of IFN-γ and IL-12, representative Th1 cytokines, were increased by IVIG treatment, but not by placebo. The percentage of IFN-γ- and IL-12-positive cell number/CD4-positive T lymphocyte cell number was also significantly increased by IVIG in SSc skin. Furthermore, mRNA expression of IFN-γ and IL-12 in SSc skin tissue was significantly up-regulated after IVIG treatment.ConclusionThe expression of Th1 cytokine is reported to be decreased in SSc. Our study suggested IVIG recovered the suppressed levels of Th1 cytokines, and that the treatment improves skin fibrosis by correcting the Th1/Th2 balance. In order to facilitate the clinical use of IVIG for SSc, it is necessary to perform a larger study in the future.

Laparoscopic and open splenectomy for splenomegaly secondary to liver cirrhosis: an evaluation of immunity

We sought to investigate the perioperative inflammatory response and immunological function of patients with portal hypertension-induced splenomegaly who underwent laparoscopic (LS) or open splenectomy (OS). This prospective study investigated a total of 34 patients with splenomegaly due to portal hypertension who underwent either LS (n=18) or OS (n=16) between May 2009 and September 2010. Peripheral venous blood samples were taken from these patients prior to surgery and on postoperative days (POD) 1, 3, and 7. The perioperative clinical outcomes and immunological function results were analyzed and compared within each surgical group. The demographics of the two groups did not differ. The patients in the LS group experienced longer operating time, less intraoperative blood loss, earlier resumption of diet, and shorter postoperative hospital stay. Both the open and laparoscopic groups exhibited statistically significant differences in interleukin -6 and C-reactive protein levels, and total lymphocyte, CD4 T, and natural killer cell numbers on POD 1 and 3 compared with pre splenectomy. The immune responses in the LS group were significantly lower than those in the OS group. The LS group exhibited better preserved cellular immune response and faster recovery than the OS group on POD 7. An examination of the inflammatory reaction and cellular immune response after LS and OS demonstrated that there are significant differences in the immune responses observed in the two groups. Further human studies are required to determine the permanent effects of LS on immune function.

Decoy Receptor 3 Attenuates Collagen-induced Arthritis by Modulating T Cell Activation and B Cell Expansion

To investigate the immune-modulated effects of decoy receptor 3 (DCR3) in an experimental model of rheumatoid arthritis (RA). We delivered DCR3 plasmid into collagen-induced arthritis (CIA) mice using the hydrodynamic method and evaluated the serum level of DCR3 protein by ELISA. After immunization, we assessed disease severity of arthritis incidence, arthritis scores, paw thickness, and means of arthritic limbs, and used hematoxylin and eosin staining to observe synovial hyperplasia. We analyzed numbers of murine splenocytes and inguinal lymphocyte cells, cell populations, and serum proinflammatory cytokines by flow cytometry. We investigated B cell proliferation by carboxyfluorescein succinimidyl ester assay. We evaluated serum levels of total IgG2a and type II collagen-specific IgG and IgG2a using ELISA. DCR3 expression in sera significantly attenuated disease severity in CIA mice. We found that DCR3 inhibited the volume of inguinal lymph nodes, numbers of CD19+ B cells, and populations of interferon-γ, interleukin 4 (IL-4), IL-17A, and Foxp3-producing CD4+ T cell in vivo. We found that DCR3 inhibited Pam3CSK4 (Toll-like receptor 1/2 ligand)-induced B220+ B cell proliferation in vitro. DCR3 treatment reduced the serum level of IL-6, total IgG2a, and CII-specific IgG2a antibody. We postulated that the protective effects of DCR3 in CIA resulted from modulation of the immune system by maintaining the B/T cell balance and decreasing lymphocyte expansion. We suggest DCR3 as a prophylactic and potential therapeutic agent in the treatment of RA.

Comparative studies on temperature threshold for heat shock protein 70 induction in young and adult Murrah buffaloes

To know the temperature threshold for heat shock protein 70 (HSP70) induction in lymphocytes and to assess physiological changes, if any, in relation to HSP70 induction in young and adult Murrah buffaloes, this study was divided into two parts: I. In vivo study: where assay of HSP70 was performed in blood samples collected from acutely exposed young and adult Murrah buffaloes (n = 6) inside a climatic chamber at 40, 42 and 45 °C for 4 h and thermoneutral temperature (22 °C). Physiological parameters viz., rectal temperature, respiratory rate, pulse rate and skin temperature of different body parts were monitored to assess magnitude of stress in the animals owing to thermal exposure II. For in vitro study, equal numbers of lymphocyte cells were separated from blood collected from young and adult buffaloes and were subjected to four temperature treatments (38, 40, 42 and 45 °C) for 4 h. A significant increase (p < 0.05) in all the physiological parameters in both young and adult buffaloes was observed after exposure to 40, 42 and 45 °C for 4 h as compared to 38 °C. The average plasma HSP70 concentrations (ng/ml) were significantly higher (p < 0.05) at 40, 42 and 45 °C as compared to 38 °C in both young and adult and were higher in young than adult buffaloes at 38 and 45 °C. Heat shock protein 70 level in lymphocyte lysate showed highest concentration after 3-h exposure to all temperatures (40, 42 and 45 °C) in both young and adult buffaloes. The intensity of changes of all physiological parameters was more in young animals than in the adults indicating the greater susceptibility of younger animals to heat stress and was found to be changed at around 40 °C when animals were exposed to different temperatures, indicating the possibility that HSP70 production may be initiated at this temperature which is 2 or 3 °C higher than core body temperature.