Effect of TIPE1 on Immune Function of Dendritic Cells and Its Signaling Pathway in Septic Mice.

Dendritic cell (DC) dysfunction plays a pivotal role in sepsis-induced immunosuppression. Tumor necrosis factor α (TNF-α)-induced protein 8 like-1 (TIPE1), a new member of the tumor necrosis factor α-induced protein 8 family, may be related to cell death. The aim of the present study was to elucidate the effect of TIPE1 on the immune function of DCs and its regulatory mechanism via PD-L1/PD-1 signaling in mice. Sepsis was induced in adult C57BL/6 male mice via cecal ligation and puncture. In vitro, we found that expression of CD80, CD86, and major histocompatibility complex class II in DCs and levels of cytokines, including tumor necrosis factor α and interleukin 12p40, were elevated; similarly, T-cell proliferation and differentiation were promoted when the gene expressing TIPE1 was silenced. Next, we examined the in vivo role of TIPE1 in a cecal ligation and puncture animal model system. Flow cytometry of the immune functional status in DCs revealed negative regulation of TIPE1 on DC maturation, as well as activation. Moreover, changes in PD-L1/PD-1 levels confirmed the negative effect of TIPE1 in DCs. Collectively, we report that TIPE1 might exert negative regulation in sepsis, at least in part by inhibiting DC maturation and subsequent T-cell-mediated immunity via PD-L1/PD-1 signaling.

Adoptive transfer of bone marrow-derived dendritic cells decreases inhibitory and regulatory T-cell differentiation and improves survival in murine polymicrobial sepsis.

A decrease in the number of dendritic cells (DCs) is a major cause of post-sepsis immunosuppression and opportunistic infection and is closely associated with poor prognosis. Increasing the number of DCs to replenish their numbers post sepsis can improve the condition. This therapeutic approach could improve recovery after sepsis. Eighty C57BL/6 mice were subjected to sham or caecal ligation and puncture (CLP) surgery. Mice were divided into four groups: (i) Sham + vehicle, (ii) Sham + DC, (iii) CLP + vehicle, and (iv) CLP + DC. Bone-marrow-derived DCs (BMDCs) were administered at 6, 12 and 24 hr after surgery. After 3 days, we assessed serum indices of organ function (alanine aminotransferase, aspartate aminotransferase, creatinine, amylase and lipase), organ tissue histopathology (haematoxylin and eosin staining), cytokine [interferon-γ (IFN-γ), tumour necrosis factor-α, interleukin-12p70 (IL-12p70), IL-6 and IL-10] levels in the serum, programmed death-1 (PD-1) expression on T cells, regulatory T-cell differentiation in the spleen, and the survival rate (monitored for 7 days). BMDC transfer resulted in the following changes: a significant reduction in damage to the liver, kidney and pancreas in the CLP-septic mice as well as in the pathological changes seen in the liver, lung, small intestine and pancreas; significantly elevated levels of the T helper type 1 (Th1) cytokines IFN-γ and IL-12p70 in the serum; decreased levels of the Th2 cytokines IL-6 and IL-10 in the serum; reduced expression of PD-1 molecules on CD4(+) T cells; reduced the proliferation and differentiation of splenic suppressor T cells and CD4(+)  CD25(+)  Foxp3(+) regulatory T cells, and a significant increase in the survival rate of the septic animals. These results show that administration of BMDCs may have modulated the differentiation and immune function of T cells and contributed to alleviate immunosuppression, hence reducing organ damage and mortality post sepsis. Hence, the immunoregulatory effect of BMDC treatment has potential for the treatment of sepsis.

N-acetylcysteine administration is associated with reduced activation of NF-kB and preserves lung dendritic cells function in a zymosan-induced generalized inflammation model.

PURPOSE: In severe sepsis, functional impairment and decreased numbers of dendritic cells (DCs) are essential reasons for immune function paralysis, secondary organ infection, and organ failure. We investigated the effects of N-acetylcysteine (NAC) administration on protecting lung DCs function in a zymosan-induced generalized inflammation (ZIGI) model. METHODS: ZIGI was initiated in 80 Balb/c mice by intraperitoneal injection of zymosan (ZYM; 900 mg/kg). Mice were divided into 4 groups: (1) SHAM+Vehicle; (2) SHAM+NAC; (3) ZYM+Vehicle; and (4) ZYM+NAC. NAC (100 mg/kg) was administered at different time after ZYM injection. After 48 h, we assessed: lung tissue pathological changes; arterial blood gas values; purified lung DCs surface expressions of MHC-II/I-A(d) and co-stimulatory molecules CD80, CD83, and CD86; lung DCs mRNA levels of chemokine receptors CCR5 and CCR7; lung DCs apoptosis; lung DCs ultrastructure by transmission electron microscopy; lung DCs NF-kB transcription factor activity; and LPS-stimulated lung DCs in vitro production of IL-12 and IL-10 were examined. RESULTS: NAC treatment resulted in: significant improvements in ZYM-induced lung tissue damage and impaired lung function; inhibited lung DCs ZYM-induced increased expression of MHC-II/I-A(d), CD83, and CD86, but not CD80; reduced lung DCs ZYM-induced CCR5 and CCR7 mRNA levels; suppressed ZYM-induced lung DCs apoptosis; ameliorated ZYM-induced lung DCs ultrastructural abnormalities; inhibited ZYM-induced lung DCs NF-κB activity; and enhanced lung DCs production of IL-12 and inhibited their production of IL-10. CONCLUSIONS: Repeated injections of NAC during the early stage of severe sepsis effectively inhibited lung DCs activation and their apoptosis, which could preserve DCs function.

[Expression of ubiquitin in the spleen and its influence on the dendritic cell immunomodulation function in multiple organ dysfunction syndrome in mice].

OBJECTIVE: To explore the expression change in ubiquitin (Ub) in the spleen and its significance in multiple organ dysfunction syndrome (MODS) in mice, and to study the effects of ubiquitination of major histocompatibility complex II (MHCII) on the activity and immunomodulation function of splenic dendritic cell (DC). METHODS: Two hundred and ten mice were divided into the normal control group (n = 30) and MODS group (n = 180) according to the method of random digital table, and MODS model was replicated by intraperitoneal injection of zymosan. The MODS group mice were further divided evenly into 6, 12, 24, 48-hour and 5-7-day and 10-12-day groups. Ub protein and expression of CD11c⁺DC and MHCII molecule I-A(b) were examined using immunohistochemistry and immunofluorescence methods. Ub mRNA expression in the spleen was measured by real-time quantitative reverse transcription--polymerase chain reaction (RT-PCR). RESULTS: (1)Immunohistochemistry results showed: the number of Ub positive cells in the spleen increased significantly at 6 hours in MODS group compared with that of the normal control group, and it reached the peak at 24 hours [(16.83 ± 0.38)% vs. (8.60 ± 0.86)%, P < 0.05] and then decreased gradually. At 10-12 days, the number of Ub positive cells decreased significantly compared with that of the normal control group [(4.66 ± 0.34)% vs. (8.60 ± 0.86)%, P < 0.05]. (2)RT-PCR results displayed: compared with normal control group, Ub mRNA expression in spleen increased at 6 hours in MODS group, and it reached the peak at 24 hours (2.17 ± 0.20 vs. 1.00 ± 0.00, P < 0.01). Then, it decreased gradually. At 10-12 days, Ub mRNA decreased significantly as compared with that of normal control group (0.72 ± 0.08 vs. 1.00 ± 0.00, P < 0.05). (3)Immunofluorescence results displayed: compared with normal control group, CD11c⁺DC increased significantly at 6 hours in MODS group and reached the peak at 24 hours [(7.55 ± 0.04)% vs. (2.08 ± 0.13)%, P < 0.05], and then it decreased gradually. At 10-12 days, it was close to that of the normal control group [(2.28 ± 0.06)% vs. (2.08 ± 0.13)%, P>0.05]. Compared with the normal control group, I-A(b) positive cells in the spleen was significantly increased at 6 hours in MODS group [(10.90 ± 1.40)% vs. (5.78 ± 0.47)%, P < 0.01], but it decreased at 24 hours [(3.32 ± 0.91)% vs. (5.78 ± 0.47)%, P < 0.05]. I-A(b) positive cells were restored to the normal level at 48 hours and 5--7 days, and decreased significantly again at 10-12 days [(2.20 ± 0.97)% vs. (5.78 ± 0.47)%, P < 0.05]. The number of Ub positive cells correlated positively to the expression of I-A(b) and the CD11c [r1 = 0.899, r2=0.987, both P < 0.05]. CONCLUSIONS: Ub might influence the maturation and activation of DC via ubiquitination of the MHCII molecule on DC, thereby influencing the immune response at different stages of MODS. The result might provide a new way to recognize immune response and also a new monitoring index for immune response regulation.

[Advanced dermatofibrosarcoma protuberans treated with imatinib mesylate].

OBJECTIVE: To evaluate the efficacy, side effects and toxicity of imatinib mesylate in the treatment of patients with locally advanced and/or metastatic dermatofibrosarcoma protuberans (DFSP). METHODS: Twenty-four cases of advanced DFSP diagnosed by pathology and treated in our hospital from Nov. 2004 to Oct. 2009 were included in this study. The patients were treated with imatinib mesylate (dosage: 400 mg, po, qd) and carefully observed for treatment efficacy, side effects and survival time. There were 2 patients taking the drug as second line therapy, and other 22 patients as third or more than third line therapy. RESULTS: The 24 patients were evaluable for the efficacy. There were 8 patients (33.3%) with CR, 10 pts (41.7%) PR, 2 patients (8.3%) SD, and 4 patients (16.7%) PD. The disease control rate (DCR = CR+PR+SD) was 83.3%. The median response time in 18 cases with CR and PR was 5.6 months. The median survival time in 20 cases with disease control was 30 months, however, that in nonresponse (PD) cases was only 10 months. Side reactions related to imatinib mesylate included nausea and vomiting (20.8%), neutropenia (12.5%), and edema (8.3%). CONCLUSIONS: Our results are consistent with previous reports in the literature. Imatinib is a safe and effective moleucular target drug used for Chinese. Only mild adverse reactions occur in the treated patients. It is worth using imatinib in the treatment of advanced DFSP patients.

[The study of follow up of percutaneous microwave ablation for uterine fibroids treatment].

OBJECTIVE: To study the prognosis of fibroid after ultrasound-guidance percutaneous microwave ablation (PMAUF). METHODS: From Mar. 2007 to Jul.2010 forty uterine fibroids in forty patients with symptoms which were diagnosed in our hospital accepted PMAUF. One day after treatment blood supply within the fibroid was evaluated, by enhanced MRI. The size of fibroid was measured by ultrasonography in one year with 3 months interval, the monthly menstrual of patient was followed and the mass discharge through vagina were collected for pathological examination. RESULTS: Among the 40 fibroid nodules, 22 are intramural fibroids, 15 subserosal fibroids and 3 submucosal fibroids. The baseline mean diameter of the fibroids ranged from (3.7 to 9.0) cm, with an average of (6.4 ± 1.5) cm. The mean volume ranged from (14.6 - 341.1) cm(3), with an average of (140.1 ± 87.4) cm(3). Three months after treatment, the anechoic zone was observed within the ablated nodules and disappeared in six to seven months. the echo of ablated zone keep higher than the surrounding tissue, whereas the size of fibroid shrank significantly. Meat tissue was discharged from vagina in 8 patients (2 cases of submucosal fibroids, 6 cases intramural fibroids) in the period of menstrual in 1 - 8 months after ablation. The largest dimension of the discharge was 2.1 cm. Pathological examination confirmed the discharges as necrotic fibroid. The patients with subserosal fibroid had no discharge of necrotic tissue. The fibroid shrink rate was higher in the patients with vaginal discharges than that without vaginal discharges (P < 0.01). CONCLUSIONS: After PMAUF necrotic tissue of submucosal and intramural fibroids can be discharged through vagina, liquefaction in the center of subserosal and intramural fibroids may occur and be absorbed gradually, that may be the main reasons for fibroids reduced significantly or disappearance.

[Changes in peripheral dendritic cells in serious burn patients and its clinical significance].

OBJECTIVE: To explore the changes in peripheral dendritic cells (DCs) in serious burn patients and its relationship with the burn severity and pathogenesis of sepsis. METHODS: Twenty-two serious burn patients were divided into the burn group (n = 10) and the burn sepsis group (n = 12) according to diagnostic criteria of sepsis, they were stratified according to the total burn surface area (TBSA), into the TBSA I group (TBSA 30%-50%, n = 14) and the TBSA II group (TBSA 51%-80%, n = 8). Peripheral blood of all patients was collected on 1, 3, 7 ,14 ,20 day after burn. The number of two subtypes of peripheral DC i. e myeloid dendritic cells [mDC, Lineage1(-)HLA-DR(+)CD11c(+)] and plasmacytoid dendritic cells [pDC, Lineage1(-)HLA-DR(+)CD123(+)] were quantified by flow cytometer. Ten healthy volunteers served as normal controls at the same time. RESULTS: In the healthy control group, mDC in the peripheral blood was (0.450 ± 0.150)% and pDC was (0.241 ± 0.084)%. Compared with the healthy control group, in the burn group both mDC [(0.257 ± 0.116)%, (0.274 ± 0.086)%, (0.317 ± 0.056)%] and pDC [(0.122 ± 0.058)%, (0.165 ± 0.051)%, (0.177 ± 0.024)%] decreased significantly on 1, 3, 7 day after burn (all P < 0.05), and the number returned to the normal level on 14 day and 20 day. Compared with the burn group, the number of mDC [(0.230 ± 0.090)%] and pDC [(0.114 ± 0.071)%] in patients of the burn sepsis group were significantly lower (both P < 0.05) on 1 day after burn. Both cells [mDC (0.246 ± 0.076)% vs. (0.412 ± 0.097)% and pDC (0.097 ± 0.032)% vs. (0.203 ± 0.039)%] were still significantly lower (both P < 0.05) on 20 day. Both mDC [(0.266 ± 0.062)%, (0.289 ± 0.071)%, (0.351 ± 0.054)%] and pDC [(0.131 ± 0.025)%, (0.163 ± 0.037)%, (0.178 ± 0.038)%] in the patients in the TBSA I group decreased significantly on 1, 3, 7 day after burn as compared with those of the healthy control group(all P < 0.05), and they returned to the normal level on 14 day and 20 day. Compared with the TBSA I group, mDC [(0.227 ± 0.070)%] and pDC [(0.112 ± 0.047)%] in patients of the TBSA II group decreased significantly(both P < 0.05)on 1 day after burn, and both cells [mDC (0.297 ± 0.072)% vs. (0.423 ± 0.046)% and pDC (0.107 ± 0.061)% vs.(0.197 ± 0.042)%] were still significantly lower (both P < 0.05) on 20 day. CONCLUSION: Both the number of mDC and pDC decrease in peripheral blood in early stage in serious burn patients, and those who have more serious burn have lower number of mDC or pDC. Deficiency in mDCs and pDC subsets may contribute to immunosuppression in burn victims, and those who suffered obvious loss of mDC and pDC are susceptible to sepsis following severe burn. It indicates that the percentage of mDC and pDC can be a predictive index of sepsis after burn.

Relationship between HMGB1 content and MHC-II expression in circulating monocytes and spleen of mice challenged with zymosan.

OBJECTIVE: To observe the regularity of change in high mobility group protein box 1 (HMGB1) content in serum and spleen of mice with multiple organ dysfunction syndrome (MODS), to analyze the correlation between HMGB1 content and major histocompatibility complex (MHC)-II---I-A(b) expression on monocytes in blood and spleen, and to explore the effect of HMGB1 on immune function of circulating monocytes and splenocytes. METHODS: One hundred 8-week-old male 57BL/6 mice were randomly divided into normal group and experimental group subdivided into 8 subgroups: 3, 8, 12 hours, 1, 2, 3, 5-7 days and 10-12 days post zymosan injection (PZI). MODS model was replicated by injecting zymosan into the peritoneal cavity. At each time point, blood and spleen were collected to detect HMGB1 content and the rate of I-A(b) positive monocytes. RESULTS: In normal and PZI 3-hour, 8-hour mice, serum HMGB1 was not detected, but it significantly increased at PZI 12 hours. In spleen of normal mice, there was low level of HMGB1 expression. In zymosan-treated mice, HMGB1 started to rise in spleen at PZI 3 hours. Subsequently, HMGB1 content in both serum and spleen significantly increased, and it reached the peak level in 1-2 days, decreased in 5 days, and then increased in 10-12 days. The number of I-A(b) positive monocytes in circulating blood and spleen decreased at 1-2 days (t equal to 9.589, 4.432, P <0.01) and 10-12 days following the challenge, forming a two trough like decrease, just corresponding with two-peak increase of HMGB1. However, at 3 hours after zymosan challenge, I-A(b) expression on circulating monocytes was downregulated (t =5.977, P less than 0.01), while that in spleen upregulated (t equal to 4.814, P less than 0.01). CONCLUSION: In mice with MODS, up-regulated HMGB1 expression can regulate I-A(b)expression on monocytes to depress their ability of presenting antigen, which results in immune disturbance contributing development of MODS.

[Effect of change in high mobility group protein box 1 expression on activity of immunocytes in spleen of mice with multiple organ dysfunction syndrome].

OBJECTIVE: To explore the regularity of high mobility group protein box 1 (HMGB1) expression and major histocompatibility complex II I-A(b) in spleen of mice with multiple organ dysfunction syndrome (MODS) , and its effect on the activity of immunocytes and relationship with pathogenesis of MODS. METHODS: MODS model was replicated by injecting zymosan into abdominal cavity of mice. The mice were randomly divided into normal control group, and 3, 8, 12 hours, and 1, 2, 3, 5, 10-12 days after injection groups. The expression levels of HMGB1, I-A(b) and apoptosis rate in the spleen were detected. RESULTS: There was a little HMGB1 expression in the spleen of control mice. After zymosan administration, HMGB1 expression was increased, it reached the highest level on 1-2 days (P<0.01), decreased on day 5, then elevated on 10-12 days (P<0.05). Change in HMGB1 mRNA expression was in accordance with that of the protein. At 8 hours following injury, the I-A(b) expressed on the splenocytes was enhanced similar to that of HMGB1, and then it reversed. The occurrence of splenocyte apoptosis was also consistent with change in HMGB1 content in the spleen. CONCLUSION: The increased apoptosis of splenocytes in mice with MODS is closely related with up-regulation of HMGB1 expression, which inhibits the expression level of I-A(b) on antigen presenting cells, thus weakens the capability of immune responses and affected the development of MODS.

[Roles of cyclooxygenase 2/2', 3'-cyclic nucleotide3' phosphohydrolase in the oligodendrocyte apoptosis in heroin-induced spongiform leucoencephalopathy].

OBJECTIVE: To investigate the regulation of cyclooxygenase (Cox)-2/2', 3'-cyclic nucleotide3' phosphohydrolase (CNPase) on the oligodendrocyte apoptosis in the pathogenesis of the heroin-induced spongiform leucoencephalopathy (HSLE). METHODS: Samples of frontal lobe, cerebellum, and corpus callosum were obtained from the brains during autopsy of 4 HSLE patients and 5 patients who died of diseases other than cerebral diseases (controls) and underwent light microscopy and electron microscopy. Immunocytochemistry was carried out to detect the expression of myelin basic protein (MBP), caspase-3, COX-2, and CNPase protein. Apoptosis was examined by TUNEL staining. RESULTS: Widespread demyelination was seen in the white matter of the frontal lobe, cerebellum, and corpus callosum of the HSLE cases, most severely in cerebellum. In he HSLE group, the levels of caspase-3 and COX-2 expression were significantly higher, and the level of CNPase was significantly lower than those of the control group (all P < 0.05). CONCLUSION: Widespread demyelination in the white matter is a prevailing pathological change of HSLE. Oligodendrocyte apoptosis is one of the causes of HSLE. The upregulation of COX-2 and downregulation of CNPase may contribute to the pathogenesis.

[Roles of bcl-2/bax expression and oligodendrocyte apoptosis in the pathogenesis of heroin-induced spongiform leucoencephalopathy].

OBJECTIVE: To investigate the role of oligodendrocyte apoptosis under the regulation of the bcl-2/bax protein expression in brain white matter in the pathogenesis of heroin-induced spongiform leucoencephalopathy (HSLE). METHODS: Samples of frontal lobe, cerebellum, and corpus callosum were obtained from the brains during autopsy of 4 HSLE cases and 5 normal controls and underwent light microscopy and electron microscopy. Immunocytochemistry was used to detect the expression of myelin basic protein (MBP), caspase-3, bcl-2 protein, and bax protein. RESULTS: Widespread demyelination was seen in the white matter of the frontal lobe, cerebellum and corpus callosum of the HSLE cases, most severely in the cerebellum. The levels of caspase-3 and bax expression of the HSLE group were significantly higher than those of the control group (both P <0.05) , however, the bcl-2 level of the HSLE group was no significantly different from that of the control group (P > 0.05). CONCLUSION: Widespread demyelination in the white matter is a prevailed pathological change of HSLE. Oligodendrocyte apoptosis under induced by the decrease of bcl-2/bax ratio may contribute to the pathogenesis.

[Flt3 ligand in recovering the function of the splenic dendritic cells and the immune system in mice with multiple organ dysfunction syndrome].

OBJECTIVE: To study the immunoprotective effect of fms-like tyrosine kinase receptor 3 ligand (FL) in multiple organ dysfunction syndrome(MODS) in mice. METHODS: Ninety mice were randomized into three groups: normal, MODS and MODS+FL (each n=30). The MODS model of mice was reproduced. Splenic dendritic cell (DC), I-Ab of peripheral blood mononuclear cells (PBMCs) and T cell subpopulation of peripheral blood of mice were analyzed by flow cytometry. The histomorphology of spleen and DC was studied by light microscope and electron microscope. RESULTS: In MODS group, the number of splenic immature DC increased (P<0.05), the number of CD4+ T lymphocytes reduced, while the number of CD8(+)T lymphocytes increased, thus CD4/CD8 ratio was reduced (P<0.05). White pulp of the spleen dispersed, with decrease in splenic bodies. There was apoptosis of DC. The expression of I-Ab of the mononuclear cells decreased remarkably (P<0.01). The mortality of mice before treatment was 18%. In FL treatment groups, the number of splenic mature DC was markedly increased (P<0.05). The number of CD4+ T lymphocytes and CD4/CD8 ratio was also markedly elevated. The expression of I-Ab of the mononuclear cells became normal. The pathological changes in spleen were alleviated. The mortality rate was significantly lowered (7%). CONCLUSION: FL can not only stimulate the recovery the DC activity, but also improve the immune function in the late phases of MODS.

Characteristics of and strategies for patients with severe burn-blast combined injury.

BACKGROUND: Severe burn-blast combined injury is a great challenge to medical teams for its high mortality. The aim of this study was to elucidate the clinical characteristics of the injury and to present our clinical experiences on the treatment of such cases. METHODS: Five patients with severe burn-blast combined injuries were admitted to our hospital 77 hours post-injury on June 7, 2005. The burn extent ranged from 80% to 97% (89.6% +/- 7.2%) of TBSA (full-thickness burns 75% - 92% (83.4% +/- 7.3%)). All the patients were diagnosed as having blast injury and moderate or severe inhalation injury. Functions of the heart, liver, kidney, lung, pancreas and coagulation were observed. Autopsy samples of the heart, liver, and lungs were taken from the deceased. Comprehensive measures were taken during the treatment, including protection of organ dys function, use of antibiotics, early anticoagulant treatment, early closure of burn wounds, etc. All the data were analyzed statistically with t test. RESULTS: One patient died of septic shock 23 hours after admission (four days after injury), the others survived. Dysfunction of the heart, liver, lungs, pancreas, and coagulation were found in all the patients on admission, and the functions were ameliorated after appropriate treatments. CONCLUSIONS: Burn-blast combined injury may cause multiple organ dysfunctions, especially coagulopathy. Proper judgment of patients' condition, energetic anticoagulant treatment, early closure of burn wounds, rational use of antibiotics, nutritional support, intensive insulin treatment, timely and effective support and protection of organ function are the most important contributory factors in successful treatment of burn-blast combined injuries.

[Implications of lung interstitial dendritic cells of mice in multiple organ dysfunction syndrome].

OBJECTIVE: To explore the role of lung interstitial dendritic cells in immunodissonance and organ injury in multiple organ dysfunction syndrome (MODS). METHODS: Animal model of MODS was established by injecting zymosan into the peritoneal cavity of C57BL/6 mice. The mice were randomly divided into groups of normal, 3 - 6 hours, 12 - 48 hours, 5 - 7 days, 10 - 12 days post injection. Pathological changes of lung and interstitial dendritic cells were studied by light and transmission electron microscope. Immunohistochemistry, RT-PCR and flow cytometry analyses were used to document status of biomarkers, including specific surface markers (CD205 and CD11c), costimulatory molecules (CD80 and CD86), SLC and its receptor CCR7 in lung, CD4+ and CD8+ T lymphocyte subtypes in peripheral blood. RESULTS: At early stage of injury, interstitial dendritic cells showed an increase in proliferation with expression of low level of CD80 and CD86. In contrast, the expression of SLC and its receptor CCR7 in lung were increased. The ratio of CD4+/CD8+ declined in peripheral blood. At the stage of SIRS, interstitial dendritic cells continued to proliferate with high expressions of CD80 and CD86. SLC and CCR7 in lung also increased. The ratio of CD4+/CD8+ declined markedly in peripheral blood. At the MODS stage, interstitial dendritic cells further proliferated, but the expression of CD80 and CD86 declined to a very low level. Although the level of SLC increased consistently, the level of CCR7 continued to decrease, along with a markedly decreased CD4+/CD8+ ratio in peripheral blood. CONCLUSIONS: Alterations of lung interstitial dendritic cells are likely to influence the course of immunological dysfunction of MODS. The level of CCR7 may serve as an indicator of the migration activity of interstitial dendritic cells and systemic immune response.

[Effects of carbachol on apoptosis of intestinal epithelial cells after gut ischemia/reperfusion in rat].

OBJECTIVE: To investigate the effects of carbachol on apoptosis of intestinal epithelial cells in rats after gut ischemia/reperfusion (I/R). METHODS: A jejunal sac was formed in Wistar rats. The superior mesenteric artery (SMA) was occluded for 45 minutes followed by 240 minutes of reperfusion. Immediately after occlusion of SAM blood flow, either 0.1 mg/kg of carbachol or same amount of 0.9% saline normal was injected into the jejunal sac. Animals were randomized into three groups (each n=40): sham operation, I/R+normal saline injection (I/R model) and I/R+carbachol injection (0.1 mg/kg, Ca). The Pathological changes in gut epithelial cells were assessed by Chiu's scores. The apoptosis index of intestinal epithelial cell was determined with terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL) staining. Expressions of caspase-3 and Bcl-2 in intestinal epithelial cells were assayed by immunohistochemistry method. All measurements were done at 0, 30, 60, 120 and 240 minutes after reperfusion. RESULTS: The Pathological injuries were less severe in Ca group than those in I/R model group. Apoptosis index of intestinal epithelial cell and expressions of caspase-3 were significantly decreased, while the expressions of Bcl-2 increased dramatically (all P<0.01) after I/R in Ca group compared with those in I/R model group, especially at 120 minutes after reperfusion. CONCLUSION: Enteral administration of carbachol can inhibit apoptosis of intestinal epithelial cells in rats after gut I/R injury.

[Change and role of liver interstitial dendritic cells of mice in a model of multiple organ dysfunction syndrome].

OBJECTIVE: To explore the role of liver interstitial dendritic cells in immuno-dissonance mechanism in multiple organ dysfunction syndrome (MODS). METHODS: The model of MODS was replicated by intraperitoneal zymosan injection in C57BL/6 mice. One hundred and fifty mice were randomly divided into groups of normal, 3-6 hours, 12-48 hours, 5-7 days and 10-12 days after zymosan injection. Morphological changes in liver interstitial dendritic cells were observed with light and transmission electron microscope. Specific surface markers CD205 and CD11c, costimulatory molecules CD80 and I-A(b), CD4+, CD8+ T lymphocyte subgroups and ratio of CD4+/CD8+ in peripheral blood were detected by immunohistochemistry and flow cytometry. RESULTS: In early stage of the challenge, liver interstitial dendritic cells showed a proliferation expressing markers CD80 and I-A(b) in a low level. The ratio of CD4+/CD8+ declined in peripheral blood. In acute stage (12-48 hours), interstitial dendritic cells had a continuous proliferation with high expression of CD11c, CD205, CD80 and I-A(b)(all P<0.01)and the ratio of CD4+/CD8+ also declined markedly in peripheral blood as compared with the normal group (P<0.01). In function failure stage (10-12 days), interstitial dendritic cells further proliferated, but the expression of CD205, CD80 and I-A(b) declined to a very low level, with comparison to that in acute stage (P<0.05 or P<0.01), and the ratio of CD4+/CD8+ declined remarkably in peripheral blood. CONCLUSION: Liver interstitial dendritic cells participated and influenced the course of dysfunction and suppression of immunity in MODS.

[Effect of carbachol on dendritic cell function in the lipopolysaccharides induced murine sepsis model].

OBJECTIVE: To explore the mechanism of the effect of carbachol in the treatment of sepsis at the angle of modulation of dendritic cell (DC) activity. METHODS: Thirty male C57BL/6 mice were randomized into three groups: normal control, sepsis and carbachol. The sepsis model of mice was reproduced by the injection of lipopolysaccharides (LPS). The expression of interleukin-1 beta (IL-1 beta) and IL-12 p70 positive cells were measured by immunohistochemistry method. The changes in DC activity in the spleen of mice were studied using flow cytometry. RESULTS: In LPS group, the number of splenic DC was increased, but it was not statistically significant. The expression of DC molecules including major histocompatibility complex-II (MHC-II), CD86 and the rate of positive cells containing IL-1 beta and IL-12 p70 were significantly higher than those in normal controls (all P<0.05). While in carbachol treatment group, the number of splenic DC had no significant change, the expression of IL-1 beta was down-regulated, dramatically. The expression of MHC-II, CD86 and IL-12 p70 were also lowered (all P<0.05). CONCLUSION: Carbachol can lower the DC activity, suggesting that modulation of DC activity may lower extensive immune and inflammatory response in the early phases of sepsis.

[Changes in spleen dendritic cells in multiple organ dysfunction syndrome in mice].

OBJECTIVE: To explore the pathological features of the splenic dendritic cells (DCs) and their role in the pathogenesis of multiple organ dysfunction syndrome (MODS). METHODS: The MODS model of mice was reproduced. The changes in DCs in the spleen in mice with MODS were studied using light microscope, electronic microscope, immunohistochemistry (CD205, CD86) and transferase mediated dUTP-biotin nick end labeling (TUNEL). RESULTS: In the early stage (3 hours), there was hyperplasia with increased activity of DCs in the spleen, accompanied by apoptosis of a large number of lymphocytes. CD4(+)/CD8(+) ratio declined in the peripheral blood. In the MODS stage (10-12 days), the number of DCs increased but with declined function. CD4(+)/CD8(+) ratio declined markedly in peripheral blood. CONCLUSION: During the course of MODS, changes in the number and activity of DCs in spleen are very important in the establishment of immune response and maintenance of immune function.

[Replication of animal model and pathological study of delayed multiple organ dysfunction syndrome in mice].

OBJECTIVE: To replicate a model of delayed multiple organ dysfunction syndrome (MODS) in mice. METHODS: Kunming mice of 6-8 week age were randomly divided into normal control group and experimental group. They were sacrificed on days 0 to 2, days 3 to 7, days 8 to 12. Symptoms and signs, survival rate, organ function parameters and pathological changes in main organs were observed in mice of all groups. RESULTS: Eight days after zymosan administration, animals showed MODS-like changes. Plasma level of alanine aminotransferase(ALT), blood urea nitrogen(BUN) and creatinine(Cr) were increased and severe pathological lesions were observed in tissues of the lung, liver, heart, kidney and gut, etc. CONCLUSION: A delayed model of MODS is replicated in mice by intraperitoneal injection of zymosan.

[A small dose of intraperitoneal injection of zymosan induces systemic inflammatory response and multiple organ dysfunction following gut ischemia-reperfusion injury].

OBJECTIVE: To investigate the effects of a small dose of intraperitoneal injection of zymosan following gut ischemia-reperfusion injury on systemic inflammatory response and distant organ function. METHODS: Wistar rats were randomly divided into three groups: gut ischemia-reperfusion injury (I-R) alone, intraperitoneal injection of zymosan (Z) alone, and gut ischemia-reperfusion followed by intraperitoneal injection of zymosan (M). Gut (I-R) was produced by occluding superior mesenteric artery for 60 minutes followed by gut reperfusion. A small dose(125 mg/kg) of zymosan was given intraperitoneally at 12 hours after recovery of gut blood flow. RESULTS: Systemic inflammatory responses were found in all rats of three groups. The plasma level of tumor necrosis factor-alpha and activities of myeloperoxidase in lungs and intestines were significantly higher in animals in M group compared with those in I-R and Z groups. The incidence of multiple organ dysfunction syndrome (MODS) and mortality rate at 72 hours postinjury in two-hits group were significantly higher than those in I-R and Z group. CONCLUSION: A small dose of intraperitoneal injection of zymosan could induce systemic inflammatory response and multiple organ dysfunction more readily when there is a precedent gut ischemia-reperfusion injury.