Important role for macrophages in induction of crescentic anti-GBM glomerulonephritis in WKY rats.

BACKGROUND: A crucial role for CD8(+) cells in induction of crescentic anti-glomerular basement membrane (GBM) glomerulonephritis (GN) in WKY rats was demonstrated in studies showing that depletion of CD8(+) cells completely suppressed glomerular accumulation of monocytes/macrophages (Mo/Mphi), crescent formation and proteinuria. Because these studies did not definitively identify CD8(+) cells as the cause of tissue injury, we examined the roles of Mo/Mphi in the development of anti-GBM GN. METHODS: We examined correlations between the amount of urinary protein and the numbers of glomerular CD8(+) cells or Mo/Mphi in rats after administrating different doses of anti-GBM antibody (5.0, 7.5, 10.0 and 25.0 microl/100 g body weight). The roles of Mo/Mphi in induction of GN were examined in animals by depleting Mo/Mphi in the glomerulus. To do this, rats were injected intravenously with liposome-encapsulated dichloromethylene diphosphonate (liposome-MDP) from day 3 to day 7 after anti-GBM antibody injection and they were then sacrificed at day 8. RESULTS: Liposome-MDP treatment significantly reduced the number of ED-1(+) Mo/Mphi accumulated in glomeruli from 32.1 +/- 1.2 to 1.4 +/- 0.3/glomerular cross-section (mean +/- SD, P < 0.01), and the amount of urinary protein from 103.8 +/- 19.8 to 31.8 +/- 15.9 mg/day (P < 0.01), as well as the incidence of crescentic glomeruli from 91.3 +/- 2.7 to 23.3 +/- 7.6% (P < 0.01) at day 8. This treatment also reduced the number of CD8(+) cells accumulating in the glomeruli from 5.4 +/- 0.7 to 0.5 +/- 0.1/glomerular cross-section (P < 0.01). Upregulation of glomerular intercellular adhesion molecule 1 (ICAM-1) and monocyte chemoattractant protein 1 (MCP-1) mRNA expression was suppressed by Mo/Mphi depletion. CONCLUSION: These results indicate that Mo/Mphi play an important role in the induction of crescentic anti-GBM GN and glomerular injury.     

Effect of a novel immunosuppressant, ST1959, on the immune system and renal allograft survival in rats

BACKGROUND: ST1959 is a 3,5-diaryl-s-triazole belonging to a novel class of contragestional agents with immunosuppressant activity. The aim of the present study was to investigate the effects of this drug on allogeneic immune response and on renal allograft survival in rats. METHODS: One group of naive and one group of allosensitized Lewis rats received ST1959 (0.5 mg/kg/day for 6 days administered subcutaneously). The respective control groups received vehicle alone. At the end of treatment, all rats were killed and thymus, spleen, lymph nodes, bone marrow, and blood were harvested. Cell number, leukocyte subpopulations, and lymphocyte alloreactivity were evaluated. Three additional groups of Lewis rats received an allogeneic (Brown Norway [BN]) kidney transplant: two groups received ST1959 (0.5 mg/kg daily until death or for 6 days and then twice weekly), and the last one received vehicle. RESULTS: In naive rats, ST1959 reduced the percentage of CD4CD8 (74.2+/-2.7%; vehicle, 89.1+/-1.1%; P<0.05) and increased the percentage of CD4CD8 thymocytes (5.7+/-0.8% vs. 2.8+/-0.4%; P<0.05). Infusion of allogeneic (BN) splenocytes caused a twofold increase of activated CD4 T cells (CD4CD25) that was prevented by ST1959 treatment. Consistently, the alloreactivity of lymphocytes from naive and allosensitized animals treated with ST1959 was significantly lower than that of control rats. ST1959 (in both tested regimens) significantly prolonged renal allograft survival in comparison with vehicle (12.4+/-0.5 vs. 7.7+/-0.5 days; P<0.001). CONCLUSIONS: ST1959 possesses immunomodulatory effects and significantly prolongs survival of renal allografts in rats.     

Podocyte changes upon induction of albuminuria in Thy-1.1 transgenic mice.

BACKGROUND: Thy-1.1 transgenic mice, characterized by ectopic expression of the Thy-1.1 protein on podocytes, spontaneously develop proteinuria and focal glomerulosclerosis (FGS). Injection of a monoclonal antibody (mAb) directed against the Thy-1.1 protein in young transgenic mice induces a massive albuminuria that is followed by an accelerated FGS within 3 weeks. This albuminuria is complement and leukocyte independent. The time course of proteinuria, the pathogenesis of the acute proteinuria and the dose dependency of FGS are unknown. METHODS: Albuminuria was measured in Thy-1.1 transgenic mice after injection of different doses of anti-Thy-1.1 mAb and at different time points within the first 24 h after injection. Podocytic foot processes and slit pore diameter were quantitated by electron microscopy. Changes in expression of slit pore constituents (podocin, CD2AP, nephrin and ZO-1), cytoskeleton-associated proteins (actin, alpha-actinin, ezrin and synaptopodin), the GDH-podocyte adhesion molecules alpha(3)-integrin, and heparan sulfate were studied by immunofluorescence. FGS was scored by light microscopy at 3 weeks after induction of albuminuria. RESULTS: Albuminuria in Thy-1.1 transgenic mice was observed within 10 min after anti-Thy-1.1 mAb injection. This rapid development of albuminuria was accompanied by a reduction in number of podocytic foot processes from 20.0 +/- 0.7/10 microm glomerular basement membrane (GBM) in saline-treated transgenic mice to 8.0 +/- 0.5 and 2.2 +/- 0.2 in anti-Thy-1.1-treated mice, at 10 min and 8 h after treatment, respectively. In addition, we observed a significant decrease in width of remaining slit pores, from 32.7 +/- 1.1 to 26.8 +/- 1.4 nm at 10 min after mAb injection. By immunofluorescence, we did not observe major changes in the expression pattern of any of the proteins studied. There was no correlation between the injected dose of the anti-Thy-1.1 mAb and the acute albuminuria. In contrast, the percentage of FGS at 3 weeks correlated with the dose, and a significant correlation between the percentage of FGS and the time-averaged albuminuria over the 3 week study period (P < 0.001) was found. CONCLUSION: Injection of mAb directed against the Thy-1.1 protein, in young non-albuminuric Thy-1.1 transgenic mice, induced an acute albuminuria within 10 min, which was accompanied by foot process effacement. Notably, we observed a decrease in slit pore width although the expression of slit pore proteins was unchanged. Also, the acute albuminuria could not be related to alterations in cytoskeleton-associated proteins, the GBM adhesion molecule alpha(3)-integrin or heparan sulfate in the GBM. The dose-dependent development of FGS and the correlation between the percentage FGS and time-averaged albuminuria suggest that, in our model, FGS is a consequence of podocyte injury. However, the data leave open the possibility that albuminuria itself contributes to FGS development. The Thy-1.1 transgenic mouse model is an excellent model to study further the relationship between podocytic injury, albuminuria and the development of FGS.     

Experimental autoimmune glomerulonephritis induced by homologous and isologous glomerular basement membrane in Brown-Norway rats

In order to study disease mechanisms and potential forms of therapy in glomerulonephritis, a model of experimental autoimmune glomerulonephritis (EAG) has been developed in the rat. We have examined the response of Brown-Norway (BN) rats to a single i.m. injection of collagenase-solubilised homologous (Sprague-Dawley, SD) or isologous (BN) glomerular basement membrane (GBM), with and without complete Freund's adjuvant (CFA). There was a dose-dependent circulating anti-GBM antibody response to all preparations of rat GBM. Animals given either antigen alone at a dose of 2 mg/kg developed circulating anti-GBM antibodies, which reached peak values by 6 weeks (63 +/- 5% following SD GBM; 53 +/- 8% following BN GBM), but did not develop glomerular deposits of IgG or nephritis. Animals given 2 mg/kg SD GBM in CFA developed greater concentrations of anti-GBM antibody by 6 weeks (122 +/- 20%) together with linear deposits of IgG on glomerular and tubular basement membranes (TBM), albuminuria (mean 7 mg/24 h), and variable focal segmental necrotising glomerulonephritis with mild interstitial nephritis. The same dose of BN GBM in CFA produced similar concentrations of circulating antibody (144 +/- 26%), with linear deposits of IgG on GBM but rarely TBM, little albuminuria, and variable mild focal glomerulonephritis. Other strains injected with SD GBM in CFA showed a variable circulating anti-GBM antibody response, which was similar to that of BN rats in PVG and DA rats but lower in LEW and WAG rats. Linear deposits of IgG on the GBM were detected in a proportion of PVG and DA rats, but not in LEW or WAG rats.(ABSTRACT TRUNCATED AT 250 WORDS)     

Inhibition of vasospasm with lymphocyte function-associated antigen-1 monoclonal antibody in a femoral artery model in rats

OBJECT: The authors have previously shown that a monoclonal antibody (mAb) that recognizes intercellular adhesion molecule-1 (ICAM-1), also known as CD54, when administered systemically inhibits experimental vasospasm in a rat femoral artery model, suggesting that ICAM-1 and leukocyte-endothelial adhesion play a crucial role in the molecular chain of events leading to posthemorrhagic vasospasm. In this report the authors confirm this hypothesis with mAbs directed against lymphocyte function-associated antigen-1 ([LFA-1] CD11a/CD18), the molecule on the surface of leukocytes that interacts with ICAM-1. METHODS: Femoral arteries in 38 Sprague-Dawley rats were isolated and exposed to autologous blood. Twenty-nine animals were then randomized into three groups and received intraperitoneal injections of anti-LFA-1 mAb (10 rats), anti-ICAM-1 mAb (10 rats), or an isotype-matched control mAb (nine rats). Injections were administered at 3 hours and 3, 6, and 9 days after surgery. Before their deaths, six animals underwent spleen harvest, and splenocytes were used in fluorescence-activated cell sorter (FACS) analysis to verify saturation of appropriate binding sites. Animals were killed at 12 days and vessels were harvested for histological study and measurement of the luminal cross-sectional area. Nine animals were randomized as earlier, killed 24 hours after a single injection of mAb, and evaluated for periadventitial infiltration of granulocytes and macrophages. Results of FACS analysis demonstrated saturation of both LFA-1 and ICAM-1 binding sites in animals treated with the respective mAb. The mean ratios of blood-exposed to saline-exposed luminal cross-sectional areas (expressed as the percentage of lumen patency) were 90.1 +/- 5.8% (mean +/- standard error of the mean) for animals treated with the anti-LFA-1 mAb (p = 0.0218), 94.2 +/- 3.3% for animals treated with the anti-ICAM-1 mAb (p = 0.0067), and 62 +/- 7.4% for animals treated with the isotype-matched control mAb. Macrophage and granulocyte counts in the periadventitial region were 39.5 +/- 3.2/hpf for animals treated with anti-LFA-1 mAb (p = 0.001), 42 +/- 3.7/hpf for animals treated with anti-ICAM-1 mAb (p = 0.003), and 72.2 +/- 6.2/hpf for control animals. CONCLUSIONS: The systemic administration of anti-LFA-1 or anti-ICAM-1 mAb initiated 3 hours after exposure to autologous blood inhibits the development of delayed chronic vasospasm at 12 days in a rat femoral artery model and leads to a significant reduction in periadventitial inflammatory cells at 24 hours. The authors conclude that blocking the migration of inflammatory cells across the endothelial surface of an artery after adventitial exposure to blood prevents the initiation of biological cascades necessary for the subsequent development of chronic vasospasm.     

Prolonged allograft survival in anti-CD4 antibody transgenic mice: lack of residual helper T cells compared with other CD4-deficient mice

BACKGROUND: Investigations of the role of CD4 T lymphocytes in allograft rejection and tolerance have relied on the use of mouse models with a deficiency in CD4 cells. However, in mice treated with depleting monoclonal antibody (mAb) and in MHC class II knockout (KO) mice, there are residual populations of CD4 cells. CD4 KO mice had increased CD4- CD8-TCRalphabeta+ helper T cells, and both strains of KO mice could reject skin allografts at the normal rate. In this study, transgenic mice with no peripheral CD4 cells were the recipients of skin and heart allografts. Results were compared with allograft survival in CD4 and MHC class II KO mice. METHODS: GK5 (C57BL/6 bml mice transgenic for a chimeric anti-CD4 antibody) had no peripheral CD4 cells. These mice, and CD4 and class II KO mice, received BALB/c or CBA skin or cardiac allografts. Some GK5 mice were treated with anti-CD8 mAb to investigate the role of CD8 cells in rejection. CD4 and CD8 cells were assessed by FACS and immunohistochemistry. RESULTS: BALB/c skin on GK5 mice had a mean survival time +/- SD of 24+/-6 days, compared with 9+/-2 days in wild-type mice. Anti-CD8 mAb prolonged this to 66+/-7 days. BALB/c skin survived 10+/-2 days on class II KO and 14+/-2 days on CD4 KO, both significantly less than the survival seen on GK5 recipients (P<0.001). BALB/c hearts survived >100 days in GK5 recipients and in wild-type recipients treated with anti-CD4 mAb at the time of grafting, in contrast to a mean survival time of 10+/-2 days in untreated wild-type mice. Immunohistochemistry revealed that long-term surviving heart allografts from the GK5 recipients had CD8 but no CD4 cellular infiltrate. These hearts showed evidence of transplant vasculopathy. CONCLUSIONS: The GK5 mice, with a complete absence of peripheral CD4 cells, provide the cleanest available model for investigating the role of CD4 lymphocytes in allograft rejection. Prolonged skin allograft survival in these mice compared with CD4 and MHC class II KO recipients was clearly the result of improved CD4 depletion. Nevertheless, skin allograft rejection, heart allograft infiltration, and vascular disease, mediated by CD8 cells, developed in the absence of peripheral CD4 T cells.     

CD44+-ESA+在结肠癌干细胞分选中的应用

目的 分选结肠癌细胞株SW480细胞中的CD133+-CD44+-ESA+亚群细胞,并观察其致瘤性.方法 用流式细胞仪分选SW480细胞中CD133+-CD44+-ESA+、CD133--CD44+-ESA+及CD133--CD44--ESA-亚群细胞.将这3组细胞分别接种于NOD/SCID小鼠,每组5只,观察肿瘤生长...     

Inhalation Anesthetics Induce Apoptosis in Normal Peripheral Lymphocytes In Vitro

Background : The authors hypothesized that perioperative lymphocytopenia is partially caused by apoptosis of lymphocytes induced by inhalation anesthetics. Therefore, they evaluated whether sevoflurane and isoflurane induce apoptosis of normal peripheral lymphocytes.

Methods : Normal peripheral blood mononuclear cells were exposed to sevoflurane and isoflurane, and the percentages of apoptotic lymphocytes was measured by Annexin V-fluorescein isothiocyanate-7-amino actinomycin D flow cytometry after 24 h of exposure (0.5, 1.0, and 1.5 mm) and after 6, 12, and 24 h of exposure (1.5 mm). The percentages of lymphocytes with caspase 3-like activity were also measured after 24 h of exposure (1.5 mm).

Results : The percentages of apoptotic lymhocytes were increased in a dose-dependent manner (controls: 5.1 +/- 1.4%; sevo-flurane: 7.3 +/- 1.3% [0.5 mm], 9.1 +/- 1.5% [1.0 mm], 12.6 +/- 2.1% [1.5 mm]; isoflurane: 7.5 +/- 1.6% [0.5 mm], 10.5 +/- 1.5% [1.0 mm], 16.3 +/- 2.7% [1.5 mm]) after 24 h of exposure and in a time-dependent manner (controls: 1.2 +/- 0.4% [6 h], 3.4 +/- 0.7% [12 h], 5.6 +/- 1.2% [24 h]; sevoflurane: 1.8 +/- 0.4% [6 h], 6.4 +/- 1.2% [12 h], 11.3 +/- 2.2% [24 h]; isoflurane: 2.6 +/- 0.5% [6 h], 8.8 +/- 1.5% [12 h],16.0 +/- 1.9% [24 h]) at the concentration of 1.5 mm. The percentages of lymphocytes with caspase 3-like activity were increased (controls: 10.0 +/- 1.1%; sevoflurane: 13.8 +/- 1.2%; isoflurane: 17.0 +/- 1.3%).     

ETA receptor blockade induces tubular cell proliferation and cyst growth in rats with polycystic kidney disease

Tissue concentrations of ET-1 are markedly elevated in the kidneys of Han:Sprague-Dawley (Han:SPRD) rats, a model of human autosomal dominant polycystic kidney disease (ADPKD). This study analyzed whether disease progression might be attenuated by endothelin receptor antagonists. Heterozygous Han:SPRD rats received an ETA receptor antagonist (LU 135252), a combined ETA/ETB receptor antagonist (LU 224332), or placebo for 4 mo. Glomerulosclerosis, protein excretion, and GFR remained unchanged, whereas interstitial fibrosis was enhanced by both compounds. BP was not reduced by both compounds in Han:SPRD rats. Renal blood flow (RBF) decreased in ADPKD rats treated with the ETA receptor antagonist. Long-term ETA receptor blockade furthermore increased markedly the number of renal cysts (ADPKD rats, 390 +/- 119 [cysts/kidney section +/- SD]; LU 135252-treated APKD rats, 1084 +/- 314; P < 0.001), cyst surface area (ADPKD rats, 7.97 +/- 2.04 [% of total section surface +/- SD]; LU 135252-treated ADPKD rats, 33.83 +/- 10.03; P < 0.001), and cell proliferation of tubular cells (ADPKD rats, 42.2 +/- 17.3 [BrdU-positive cells/1000 cells]; LU 135252-treated ADPKD rats, 339.4 +/- 286.9; P < 0.001). The additional blockade of the ETB receptor attenuated these effects in Han:SPRD rats. Both endothelin receptor antagonists had no effect on BP, protein excretion, GFR, and kidney morphology in Sprague-Dawley rats without renal cysts. It is concluded that ETA receptor blockade enhances tubular cell proliferation, cyst number, and size and reduces RBF in Han:SPRD rats. This is of major clinical impact because endothelin receptor antagonists are upcoming clinically used drugs.     

Effects of early and late intervention with epoetin alpha on left ventricular mass among patients with chronic kidney disease (stage 3 or 4): results of a randomized clinical trial

It is not known whether prevention of anemia among patients with chronic kidney disease would affect the development or progression of left ventricular (LV) hypertrophy. A randomized controlled trial was performed with 155 patients with chronic kidney disease (creatinine clearance, 15 to 50 ml/min), with entry hemoglobin concentrations ([Hb]) of 110 to 120 g/L (female patients) or 110 to 130 g/L (male patients). Patients were monitored for 2 yr or until they required dialysis; the patients were randomized to receive epoetin alpha as necessary to maintain [Hb] between 120 and 130 g/L (group A) or between 90 and 100 g/L (group B). [Hb] increased for group A (from 112 +/- 9 to 121 +/- 14 g/L, mean +/- SD) and decreased for group B (from 112 +/- 8 to 108 +/- 13 g/L) (P < 0.001, group A versus group B). On an intent-to-treat analysis, the changes in LV mass index for the groups during the 2-yr period were not significantly different (2.5 +/- 20 g/m(2) for group A versus 4.5 +/- 20 g/m(2) for group B, P = NS). There was no significant difference between the groups in 2-yr mean unadjusted systolic BP (141 +/- 14 versus 138 +/- 13 mmHg) or diastolic BP (80 +/- 6 versus 79 +/- 7 mmHg). The decline in renal function in 2 yr, as assessed with nuclear estimations of GFR, also did not differ significantly between the groups (8 +/- 9 versus 6 +/- 8 ml/min per 1.73 m(2)). In conclusion, maintenance of [Hb] above 120 g/L, compared with 90 to 100 g/L, had similar effects on the LV mass index and did not clearly affect the development or progression of LV hypertrophy. The maintenance of [Hb] above 100 g/L for many patients in group B might have been attributable to the relative preservation of renal function.     

Low-fat, high-carbohydrate diets improve wound healing and increase protein levels in surgically stressed rats

The specific effects of omega 3 and omega 6 polyunsaturated fatty acids on wound healing, nutrition status, or immune function are controversial. Therefore, we investigated the effects of fatty acid supplementation on wound healing and nitrogen retention in a surgically stressed rat model. Male Sprague-Dawley rats (weighing 250 g) were placed into three isocaloric, isonitrogenous feeding groups (controls [standard Vivonex]; 30% safflower oil [omega 6]; or 30% fish oil [omega 3]) for 8 days prior to receiving subcutaneous vascular graft wound cylinders in their dorsal midline. Nitrogen balance was monitored daily. Wounds healed for 10 days, animals were then euthanized, serum was drawn, and wound cylinders were harvested for analyses. The low-fat, high-carbohydrate control group had higher serum albumin levels at 10 days than either fatty acid-supplemented group (3.5 +/- 0.4 g/dL v 2.9 +/- 0.3 g/dL and 2.7 +/- 0.2 g/dL, omega 3 and omega 6, respectively; both P less than .05) and had better nitrogen balance (8.6 +/- 0.8 mg N/d v -2.6 +/- 0.9 mg N/d and 0.8 +/- 1.2 mg N/d, omega 3 and omega 6, respectively; both P less than .05). They also had better healed wounds at 10 days (450 +/- 290 micrograms 5-hydroxyproline [OHP]/cm of wound cylinder v 150 +/- 40 micrograms OHP/cm and 145 +/- 90 micrograms OHP/cm, omega 3 and omega 6, respectively). Surgically stressed rats had higher protein levels, better nitrogen balance, and improved wound healing when fed a diet high in carbohydrates and low in fat.     

Rituximab in idiopathic membranous nephropathy: a one-year prospective study

Currently available monoclonal antibodies against the B cell surface antigen CD20 have been employed to explore whether specific inhibition of B cells may help improve the outcome of idiopathic membranous nephropathy (IMN) and may avoid the side effects of steroids and immunosuppressants. This prospective, observational study evaluated the 1-yr outcome of eight IMN patients with persistent (>6 mo) urinary protein excretion > 3.5 g/24 h given four weekly infusions of the anti-CD20 antibody rituximab (375 mg/m(2)). At 3 and 12 mo, proteinuria significantly decreased from mean (+/- SD) 8.6 +/- 4.2 to 4.3 +/- 3.3 (-51%, P < 0.005) and 3.0 +/- 2.5 (-66%, P < 0.005) g/24 h, albumin fractional clearance from 2.3 +/- 2.1 to 1.2 +/- 1.7 (-47%, P < 0.05) and 0.5 +/- 0.6 (-76%, P < 0.003), and serum albumin concentration increased from 2.7 +/- 0.5 to 3.1 +/- 0.3 (+21%, P < 0.05) and 3.5 +/- 0.4 (+41%, P < 0.05) mg/dl. At 12 mo, proteinuria decreased to < or =0.5 g/24 h or < or =3.5 g/24 h in two and three patients, respectively. Proteinuria decreased in the remaining patients by 74%, 44%, and 41%, respectively. Body weight, diastolic BP, and serum cholesterol progressively decreased in parallel with an improvement of edema in all patients. Renal function stabilized (Delta1/creatinine: +0.002 +/- 0.007). CD20 B lymphocytes fell below normal ranges up to study-end. No patient had major drug-related events or major changes in other laboratory parameters. Rituximab thus promotes sustained disease remission in patients otherwise predicted to progress to ESRD, and it is safe. The long-term risk/benefit profile of this novel, disease-specific approach seems much more favorable to that of commonly employed immunosuppressive drugs.     

Epidural patient-controlled analgesia: influence of bupivacaine and hydromorphone basal infusion on pain control after cesarean delivery.

Epidural administration of hydromorphone was evaluated using a patient-controlled analgesia (PCA) delivery system in 170 healthy women undergoing elective cesarean delivery with epidural bupivacaine who were randomly assigned to one of four epidural PCA treatment groups: group I, hydromorphone alone by bolus administration; group II, hydromorphone, with a continuous (basal) infusion; group III, hydromorphone in combination with 0.08% bupivacaine by bolus administration; or group IV, hydromorphone and bupivacaine, with a concurrent infusion of both drugs. Patients in group I required significantly less opioid medication (2.1 +/- 1.1 mg [mean +/- SD]) during the first 24 h than patients in group II (3.3 +/- 1.3 mg). Similarly, patients in group III self-administered significantly less hydromorphone (2.0 +/- 1.0 mg) and bupivacaine (23.3 +/- 11.4 mg) during the first 24 h of PCA therapy, compared with patients in group IV (hydromorphone [2.7 +/- 1.1 mg] and bupivacaine [31.5 +/- 11.6 mg]). The concomitant use of a local anesthetic or basal opioid infusion with hydromorphone via epidural PCA did not decrease the number of PCA demands or delivered doses. In addition, patients in all four groups had similar pain, sedation, discomfort, fatigue, and anxiety scores. The frequency of awakening at night to self-administer analgesic medication was not decreased when a basal infusion was used.(ABSTRACT TRUNCATED AT 250 WORDS)     

CCL5对MCF-7中细胞增殖能力及CD44~+/CD24~-亚群比例的影响

目的 观察外源性趋化因子5(CCL5)对人乳腺癌MCF-7细胞增殖及CD44+/CD24-亚群比例的影响,探讨其在乳腺癌进展过程中的作用.方法 噻唑蓝(MTT)比色法检测浓度为10、50、100、500 μg/L的外源性rhCCL5(recombinant human CCL5)作用0、24、48 h后MCF-7细胞增殖水平的变化,流式细胞仪检测CD44~+/CD24~-亚群比例的变化.结果 rhCCL5作用48 h后MCF-7细胞增殖水平高于对照组(P＜0.05),并呈浓度依赖趋势;且高于作用24、0 h后细胞的增殖水平(P＜0.05).50、500μg/L rhCCL5处理MCF-7细胞后升高了细胞株中CD44~+/CD24~-亚群的比例,分别为对照组的5.68和6.05倍(P＜0.05).结论 外源性CCL5可以明显升高MCF-7细胞的增殖水平以及CD44~+/CD24~-亚群的比例,这种作用可能是其诱导乳腺癌进展的机制之一.     

青春期前己烯雌酚暴露对SD大鼠睾丸发育及功能的影响

目的:研究青春期前己烯雌酚(DES)暴露对SD大鼠睾丸发育及功能的影响。方法:21日龄雄性SD大鼠90只,随机分为DES0.01、0.1、1.0、10.0μg/(kg.d)4个实验组和1个对照组(分别为Da、Db、Dc、Dd和C组,每组n=18)。于青春期前,即出生后第22d(postnatalday22,PND22)～35d(PND35),实验组每日皮下注射相应剂量的DES,共14d,对照组仅注射溶媒(玉米油)。观察各组大鼠睾丸下降时间。于青春期晚期(PND50)、性成熟后(PND64)和成年期(PND130)分3批(每批n=6)处死各组大鼠取材,测定睾丸重量,观察比较睾丸组织形态学变化,分析PND130大鼠附睾尾精子质量。结果:C、Da、Db、Dc和Dd组睾丸下降时间分别为PND26.17±1.94、26.83±1.47、28.68±1.03、33.50±1.87和41.50±2.74,其中Db、Dc和Dd组较C组明显延迟(P<0.05或P<0.01)。PND50时,C、Da、Db、Dc和Dd组单侧睾丸重量分别为(1.38±0.10)、(1.38±0.12)、(1.30±0.14)、(0.86±0.18)g和(0.73±0.27)g,其中Dc和Dd组较C组显著减轻(P<0.01);与C组比较,Db组仅有少数生精小管生精上皮中的细胞数目稍减少,Dc和Dd组生精小管发育较差、生精上皮中细胞数目减少、精子发生阻滞、间质细胞发育幼稚,其程度随DES暴露剂量增加而加重。PND64时,C、Da、Db、Dc和Dd组单侧睾丸重量分别为(1.60±0.06)、(1.62±0.11)、(1.58±0.08)、(1.47±0.10)g和(0.99±0.37)g,其中Dc和Dd组较C组显著减轻(P<0.05或P<0.01);Dc和Dd组睾丸组织形态学改变与PND50时类似,但总体较PND50有所改善。PND130时,各实验组与对照组比较,单侧睾丸重量差异无统计学意义(P>0.05),睾丸组织形态学改变难以鉴别出明显差异;C、Da、Db、Dc和Dd组大鼠附睾尾精子密度分别为(73.00±16.90)、(68.00±19.67)、(68.67±12.15)、(35.17±15.64)×106/ml和(19.13±5.17)×106/ml,其中Dc和Dd组精子密度较C组明显降低(P<0.01);与C组比较,Dd组精子活动率下降(P<0.01),Db、Dc和Dd组a级精子比例降低(P<0.05或P<0.01),Dd组b级精子比例降低(P<0.01)。结论:青春期前小剂量DES[0.01μg/(kg.d)×14d]暴露对SD大鼠睾丸发育及功能无明显影响,较大剂量DES[1.0～10.0μg/(kg.d)×14d]暴露对大鼠睾丸发育及功能有明显的近期(PND50和PND64)和较远期(PND130)毒性作用,该毒性作用随DES的暴露剂量增加而加重,随鼠龄增长而逐渐减退,其机制可能与间质细胞和支持细胞的发育及功能受损相关。     

Depletion of CD8 positive cells in nephrotoxic serum nephritis of WKY rats.

Following a small dose of nephrotoxic serum (NTS) WKY rats demonstrated crescentic glomerulonephritis, which was characterized by the early infiltration of CD8 positive cells in glomeruli. In vivo depletion of CD8 positive cells from WKY rats completely prevented proteinuria (4.6 +/- 4.8 mg/day vs. 105.3 +/- 11.6 mg/day on day 10; N = 19, P less than 0.001) and crescent formation (2.7 +/- 2.9% vs. 94.3 +/- 2.6%; P less than 0.001). Immunofluorescence revealed complete inhibition of the influx of CD8 positive cells and subsequent reduction of the infiltration of macrophages in the glomeruli. Glomerular binding of 125I-anti-rat glomerular basement membrane antibodies, host anti-rabbit IgG production and the C3 level in the circulation were the same as in the control. These data indicate that CD8 positive cells play a key role in glomerular injury and crescent formation. This model provides a useful system for studying the cellular mechanisms that lead to glomerular injury and subsequent crescent formation.     

Normalization of skeletal muscle glycogen synthesis and glycolysis in rosiglitazone-treated Zucker fatty rats: an in vivo nuclear magnetic resonance study

The aim of this study was to characterize insulin-stimulated skeletal muscle glucose metabolism in Zucker fatty rats and to provide insight into the therapeutic mechanism by which rosiglitazone increases insulin-stimulated glucose disposal in these rats. Metabolic parameters were measured using combined in vivo (13)C nuclear magnetic resonance (NMR) spectroscopy to measure skeletal muscle glucose uptake and its distributed fluxes (glycogen synthesis and glycolysis), and (31)P NMR was used to measure simultaneous changes in glucose-6-phosphate (G-6-P) during a euglycemic-hyperinsulinemic clamp in awake Zucker fatty rats. Three groups of Zucker fatty rats (fatty rosiglitazone [FRSG], fatty control [FC], lean control [LC]) were treated for 7 days before the experiment (3 mg/kg rosiglitazone or vehicle via oral gavage). Rates of glycolysis and glycogen synthesis were assessed after treatment by monitoring 1,6-(13)C(2) glucose label incorporation into 1-(13)C glycogen, 3-(13)C lactate, and 3-(13)C alanine during a euglycemic ( approximately 7-8 mmol/l)-hyperinsulinemic (10 mU. kg(-1). min(-1)) clamp. The FRSG group exhibited a significant increase in insulin sensitivity, reflected by an increased whole-body glucose disposal rate during the clamp (24.4 +/- 1.9 vs. 17.6 +/- 1.4 and 33.2 +/- 2.0 mg. kg(-1). min(-1) in FRSG vs. FC [P < 0.05] and LC [P < 0.01] groups, respectively). The increased insulin-stimulated glucose disposal in the FRSG group was associated with a normalization of the glycolytic flux (52.9 +/- 9.1) to LC (56.2 +/- 16.6) versus FC (18.8 +/- 8.6 nmol. g(-1). min(-1), P < 0.02) and glycogen synthesis flux (56.3 +/- 11.5) to LC (75.2 +/- 15.3) versus FC (16.6 +/- 12.8 nmol. g(-1). min(-1), P < 0.05). [G-6-P] increased in the FRSG and LC groups versus baseline during the clamp (13.0 +/- 11.1 and 16.9 +/- 5.8%, respectively), whereas [G-6-P] in the FC group decreased (-23.3 +/- 13.4%, P < 0.05). There were no differences between groups in intramyocellular glucose, as measured by biochemical assay. These data suggest that the increased insulin-stimulated glucose disposal in muscle after rosiglitazone treatment can be attributed to a normalization of glucose transport and metabolism.     

Infectious tolerance mediated by CD8+ T-suppresor cells after UV-B-irradiated donor-specific transfusion and rat heart transplantation

AIMS: CD8+CD28- human T-suppressor cells (Ts), which can be generated in vitro, act directly on APC rendering them tolerogenic to unprimed and primed CD4+ T cells. The aim of this study was to investigate the possibility that CD8+ T cells mediate the induction of tolerance in a heart transplantation model in rodents. MATERIALS AND METHODS: Blood from Lewis rats was UV-B-irradiated and transfused into ACI recipients on days -21, -14, and -7 before heart allograft transplantation on day 0. CD4(+) and CD8(+) T cells were positively selected from ACI rats, which had tolerated Lewis heart allografts for more than 100 days and were adoptively transferred to naive ACI rats pretreated (day -1) with gamma irradiation. These ACI rats underwent transplantation with Lewis hearts 24 hours after adoptive transfer of putative T-suppressor cells. RESULTS: Adoptive transfer of CD8(+) T cells from tolerant ACI to naive ACI rats significantly prolonged Lewis heart mean allograft survival time (MST +/- SD) to 69 +/- 13 days as compared with 15 +/- 1 and 14 +/- 1 days in animals adoptively transferred with CD4+ T cells or untreated controls, respectively (P < .001). Similarly, adoptive transfer of CD8(+) T cells from secondary ACI recipients to naive syngeneic animals also significantly prolonged survival of heart allografts to MST +/- SD of 72 +/- 4 for CD8(+) and 15 +/- 4 days for CD4(+) T cells (P < .001). CONCLUSIONS: These data demonstrate that allogeneic tolerance induced in ACI recipients by treatment with UV-B-irradiated blood from Lewis donors is mediated by CD8+ T-suppressor cells.