Hyperhomocysteinaemia therapy in haemodialysis patients: folinic versus folic acid in combination with vitamin B6 and B12.

BACKGROUND: In a recent uncontrolled retrospective report we suggested that the long-term supplementation of high-dose, i.v. folinic acid combined with high-dose i.v. pyridoxine was highly effective in correcting plasma total homocysteine (tHcy) concentrations in haemodialysis patients. To confirm these findings, we conducted a randomized, controlled trial aimed at evaluating whether i.v. or oral folinic acid provided improved tHcy-lowering efficacy in haemodialysis patients compared with oral folic acid. METHODS: In a 6-month prospective, randomized, controlled trial, 60 chronic haemodialysis patients, matched for age, gender, dialysis duration, and average screening pre-treatment-fasting tHcy levels, were given either 50 mg/week of i.v. calcium folinate (group 1), 50 mg/week of oral calcium folinate (group 2), or 45 mg/week oral folic acid (group 3). All 60 patients also received 750 mg/week of i.v. vitamin B6 and 3 mg/week of oral vitamin B12. RESULTS: Fasting tHcy decreased significantly and to a similar extent in the three groups after 2 months of treatment and remained stable at 4 and 6 months (16.6+/-3.5, 18.3+/-4, and 19.1+/-3.1, in groups 1, 2, and 3, respectively, P=NS). Mean percentage reduction at 6 months was also similar in the three treatment groups (46, 43, and 42% in groups 1, 2, and 3, respectively, P=NS). CONCLUSIONS: These findings show that the tHcy-lowering effects of high-dose i.v. folinic acid, oral folinic acid, or oral folic acid were comparable, suggesting that the hyperhomocysteinaemia observed in haemodialysis patients is not due to abnormal folate metabolism. Furthermore, they are compatible with the view that other abnormalities are also involved in the impaired clearance of homocysteine in uraemic patients.     

Linkage between O6-methylguanine-DNA methyltransferase (O6-MT) activity and cellular resistance to antitumour nitrosoureas in cultured rat brain tumour cell strains

Summary We have examined O⁶-methylguanine-DNA methyltransferase (O⁶-MT) activity of rat brain tumour cell strains with reference to cellular resistance to antitumour nitrosoureas, 1-(4-amino-2-methyl-5-pyrimidinyl) methyl-3-(2-chloroethyl)-3-nitrosourea hydrochloride (nimustine, ACNU) and methyl-6-[3-(2-chloroethyl)-3-nitrosoureido]-6-deoxy--D-glucopyranoside (ramustine, MCNU). The values of O⁶-MT activity were 52 and 160 fmol/mg protein extract in 9 L and C 6 rat brain tumour cells, respectively; while HeLa S 3 cells, as a methyl excision repair positive (Mer⁺) cell strain, revealed a rather high value of 488 fmol/mg. 9 L cells indicative of a low O⁶-MT activity showed 13 M for ACNU and 18 M for MCNU at a 10% survival dose (SD₁₀), determined by a clonogenic cell assay as an index of cellular resistance. In contrast to this, C 6 cells revealed a SD₁₀ value of 67 M and 36 M for ACNU and MCNU, respectively, indicating higher resistance than 9 L cells. HeLa S 3 cells showed the highest SD₁₀ value as follows: 84 M for ACNU and 73 M for MCNU. The relationship between the O⁶-MT activity and the cellular resistance was almost linear, with relatively resistant cell lines exhibiting the higher levels of the O⁶-MT activity. This correlation between the O⁶-MT activity and the cellular resistance to nitrosoureas as ACNU and MCNU was not observed among other antitumour drugs, which included bleomycin (BUM), neocarzinostatin (NCS),cis-diamminedichloroplatinum (II) (CDDP), and etoposide (VP-16) in clinical use for brain tumour chemotherapy. This indicates that O⁶-MT activity can be an indicator of cellular resistance to antitumour nitrosoureas in the chemotherapy of brain tumours.     

DGAT1-deficiency affects the cellular distribution of hepatic retinoid and attenuates the progression of CCl4-induced liver fibrosis

Background

Diacylglycerol O-acyltransferase 1 (DGAT1) catalyzes the final step of triglyceride synthesis, transferring an acyl group from acyl-CoA to diacylglycerol. DGAT1 also catalyzes the acyl-CoA-dependent formation of retinyl esters in vitro and in mouse intestine and skin. Although DGAT1 is expressed in both hepatocytes and hepatic stellate cells (HSCs), we reported genetic and nutritional studies that established that DGAT1 does not contribute to retinyl ester formation in the liver.

Methods

We now have explored in more depth the role(s) of DGAT1 in hepatic retinoid metabolism and storage.

Results

Our data show that DGAT1 affects the cellular distribution between hepatocytes and HSCs of stored and newly absorbed dietary retinol. For livers of Dgat1-deficient mice, a greater percentage of stored retinyl ester is present in HSCs at the expense of hepatocytes. This is also true for newly absorbed oral [³H]retinol. These differences are associated with significantly increased expression, by 2.8-fold, of cellular retinol-binding protein, type I (RBP1) in freshly isolated HSCs from Dgat1-deficient mice, raising the possibility that RBP1, which contributes to retinol uptake into cells and retinyl ester synthesis, accounts for the differences. We further show that the retinyl ester-containing lipid droplets in HSCs are affected in Dgat1-null mice, being fewer in number but, on average, larger than in wild type (WT) HSCs. Finally, we demonstrate that DGAT1 affects experimentally induced HSC activation in vivo but that this effect is independent of altered retinoic acid availability or effects on gene expression.

Conclusions

Our studies establish that DGAT1 has a role in hepatic retinoid storage and metabolism, but this does not involve direct actions of DGAT1 in retinyl ester synthesis.     

m6A对BMSCs的分化及骨质疏松症的调节作用

骨质疏松是一种全身性骨病,其特征是骨量低下、骨微结构损坏而导致骨脆性增加、易发生骨折,多见于绝经后妇女和老年男性。骨质疏松是一种退行性疾病,随年龄的增长患病风险增高。我国是世界上老年人口绝对数量最多的国家,随着人民寿命延长和老龄化社会的到来,骨质疏松已经成为我国社会的重要健康问题。N6-甲基腺苷（m6A）是位于腺苷N6位点的一种动态甲基化修饰,是真核生物mRNA中最普遍的内部修饰,介导着mRNA的剪接、结构转换、转运、翻译、降解等代谢过程。m6A由甲基化转移酶复合物催化形成,这个过程可被去甲基化酶FTO和ALKBH5逆转而m6A的“reader”蛋白通过识别m6A位点而发挥相应功能。骨髓间充质干细胞(bone mesenchymal stem cells, BMSCs)来源于骨髓,具有多向分化潜能如成骨细胞、软骨细胞、脂肪细胞等。m6A可以调节BMSCs成脂分化与成骨分化之间的转换,m6A核心的甲基化转移酶METTL3能促进BMSCs的成骨向分化,抑制其成脂向分化而FTO则相反。多个通路参与了m6A调节BMSCs成骨向分化的过程如PI3K-Akt通路、Notch通路等。m6A也调节了骨质疏松症的发生,METTL3和FTO都对骨形成有重要作用,两者可能分别介导了Ⅰ型骨质疏松症和Ⅱ型骨质疏松症。在本文中,我们总结归纳了有关m6A与BMSCs分化以及骨质疏松症的关系的最新研究成果,认为m6A可能是骨质疏松症潜在的治疗靶点。     

大鼠烧伤后库普弗细胞在促炎细胞因子产生中的作用

目的　观察大鼠严重烧伤后早期 ,库普弗细胞在肿瘤坏死因子α(TNFα)、白细胞介素(IL) 1β、IL 6产生中的作用。方法　观察 (1)烧伤血清对体外培养的大鼠库普弗细胞分泌TNFα、IL 1β、IL 6的刺激作用 ;(2 )烧伤后大鼠库普弗细胞的细胞因子mRNA表达变化 ;(3)应用库普弗细胞特异性抑制剂三氯化钆后 ,烧伤大鼠血浆内细胞因子含量变化。　结果　烧伤血清能刺激库普弗细胞释放TNFα、IL 1β、IL 6 ;大鼠烧伤后库普弗细胞TNFα、IL 1β、IL 6mRNA表达量显著升高 ;预先抑制库普弗细胞的活性 ,烧伤后血浆TNFα、IL 1β、IL 6水平均显著降低 ,分别为烧伤组的 34.71%、36 99%、33.70 %。结论　库普弗细胞是大鼠烧伤后血浆中TNFα、IL 1β、IL 6的主要来源     

结直肠癌患者手术前后血清中IL-6和NO水平的临床意义

目的探讨结直肠癌患者手术治疗前后血清中IL-6和NO水平变化及其临床意义。方法对40例结直肠癌患者分别采用酶联免疫分析法和化学比色法测定手术前后血清中的IL-6和NO水平,并与10名正常健康人作比较。结果结直肠癌患者手术前血清中IL-6水平高于正常人（P〈0.05）,NO水平明显低于正常人（P〈0.01）;经手术治疗2周后,患者血清IL-6水平较治疗前降低（P〈0.05）,而NO水平明显升高（P〈0.05）。结论结直肠癌患者血清中IL-6和NO水平的变化,对病情和预后判断具有重要的临床意义。     

急性胰腺炎合并肝损伤时IL-6、氧自由基变化及IL-2、川芎嗪干预的实验研究

目的：探讨IL－6、氧自由基在急性胰腺炎（acute pancreatitis，AP）合并肝损伤中的作用，及重组人白介素－2（IL－2）、川芎嗪的治疗价值。方法：SD大鼠112只，随机分为14只，每组8只，5％牛磺胆酸钠逆行胰胆管内注射诱发大鼠AP动物模型，检测血浆IL－6、SOD、MDA、ALT、AST、LDH、LIP、AMY，并观察肝、胰病理变化。结果；①AP组血浆AMY、LIP、ALT、AST、LDH明显升高（P＜0．05或0．01），镜下可见胰腺水肿、炎细胞浸润、坏死1肝脏肝窦充血、细胞浊肝及坏死，且损伤程度随时限延长而加重；②AP各组血浆IL－6明显升同（P＜0．01）；③AP各组MDA明显高（P＜0．01）、SOD明显降低（P＜0．01）。④IL－2治疗组、川芎嗪治疗组有IL－2、川芎嗪联合且与NS组比较血浆IL－6、MDA水平明显下降（P＜0．05），SOD明显升高（P＜0．05），胰、肝病理损害程度减轻，并且AMY、LIP、ALT、AST、LDH均明显降低（P＜0．05），平均存活时间明显延长（P＜0．05）；联合应用组降低IL－6、MDSA水平和减轻胰腺坏死优于单药组。结论：①IL－6、氧自由基在急性胰腺炎合并肝损伤程度和明显升同，起损伤作用，而SOD明显降低，其保护作用减弱。检测血浆IL－6、MDA、SOD可作为判断AP合并肝脏损伤程度和预后的指标；②大鼠急性胰腺合并肝损伤过程中应用IL－2、川芎嗪显示出良好的效果，联合应用于亿于这两药的单独应用。     

Effects of vitamin K2 and calcitonin on bone resorption model induced by vitamin A in thyroparathyroidectomized rats

Vitamin K₂ is considered to have two different effects: one is to enhance bone formation, and the other is to suppress bone resorption. However, as these effects have not been observed in a single experiment, it is unclear whether bone formation can proceed during a state of accelerating bone resorption. We therefore examined the effects of vitamin K₂ and calcitonin on a vitamin A-induced bone resorption model of thyroparathyroidectomized rats using bone histomorphometry and bone metabolism markers. The seven groups of male Sprague-Dawley rats (6 weeks old) were sham operation of thyroparathyroidectomy (TPTX) (sham group), TPTX (TPTX group), treated with vitamin A (20 mg/kg per day) from 11th to 20th day after TPTX (A group), treated with vitamin A and vitamin K₂ (30 mg/kg per day) or its vehicle from 11th to 20th day after TPTX [K group or K (veh) group], and treated with vitamin A and calcitonin (10IU/kg/ per day) or its vehicle during the same period [CT group or CT (veh) group]. Serum and urine samples were taken for marker determination on days 10, 13, 16, and 19 of TPTX and at death on the 21st day after TPTX. Undecalcified sections (Villanueva bone stain) were made of the left tibiae and decalcified sections [tartrate-resistant acid phosphatase (TRAP) stain] of the right tibiae. In the undecalcified sections, secondary trabeculae were used for histomorphometry, and in the decalcified sections primary and secondary trabeculae were used. Serum Ca of the vitamin A-administered group was significantly higher than that of the TPTX group, but this change was inhibited by vitamin K₂ or calcitonin. Serum alkaline phosphatase (ALP) in the K group was significantly higher than in all the thyroparathyroidectomized groups except the K (veh) group. In the undecalcified sections, although there was no significant difference between any of the groups in bone volume, the K group showed an increase of osteoid surface and mineralizing surface. In the decalcified sections, the K group showed a decrease of TRAP-positive areas compared to the K (veh) group in primary trabeculae. There was no significant difference between the K and K (veh) groups in secondary trabeculae. Results from the CT group were compatible with bone resorption inhibition in both bone metabolism markers and bone histomorphometry. We found that vitamin K₂ enhances bone formation and suppresses bone resorption in areas with a high turnover of bone metabolism. Vitamin K₂ is therefore expected to increase bone content if it is administered over an extended period.     

Serum and cellular interleukin-6 in haemodialysis patients: relationship with energy expenditure.

BACKGROUND: Inflammation is a highly prevalent condition among end-stage renal disease (ESRD) patients and it has been implicated with several metabolic derangements. Considering the harmful effect of hypermetabolism on nutritional status and clinical outcomes of ESRD patients, we aimed to investigate the relationship between proinflammatory cytokine interleukin-6 (IL-6) and energy expenditure in this population. METHODS: This cross-sectional study enrolled 80 adult haemodialysis patients for the evaluation of serum IL-6 and energy expenditure. The production of IL-6 by peripheral blood mononuclear cells (PBMCs) (spontaneous and endotoxin-stimulated production) was examined in a subgroup of 30 haemodialysis patients and in 11 healthy control subjects. IL-6 was measured by immunoenzymatic assay. The resting energy expenditure was evaluated by means of indirect calorimetry. Body composition was assessed by bioelectrical impedance analysis and skinfold thicknesses. RESULTS: Serum IL-6 [6.3 (2.2-163.5) pg/ml] correlated positively with age (R = 0.26; P = 0.02) and C-reactive protein (R = 0.31; P < 0.01). Resting energy expenditure correlated positively with lean body mass (R = 0.68; P < 0.001) and BMI (R = 0.44; P < 0.001), and negatively with Kt/V (R = -0.37; P < 0.01). In the multivariate analysis, controlling for age and lean body mass, serum IL-6 was positively associated with resting energy expenditure (n = 80; beta = 2.4; P = 0.01). The production of IL-6 by PBMCs did not reach statistically significant differences between patients and controls [spontaneous production 6541 (96-7739) pg/ml vs 3410 (50-7806) pg/ml, respectively; and stimulated production 6530 (579-7671) pg/ml vs 5304 (1527-7670) pg/ml, respectively]. IL-6 secreted by monocytes showed no association with either serum IL-6 or resting energy expenditure. CONCLUSION: Serum IL-6 was associated with an increase of energy expenditure in haemodialysis patients.     

Effects of 6-Chloro-6-deoxyglucose on Electrolyte and Water Transport in the Epididymis and Fertility of Male Rats

The effects of 6-chloro-6-deoxyglucose (6 CDG) on the transport of electrolytes and water in the cauda epididymidis and fertility of male rats were studied. Injection of 6 CDG into male rats at a dose rate of 120 μM/kg/day for 7–14 days induced sterility and inhibited sodium and water reabsorption in the perfused cauda epididymidis by about 60%. The rates of potassium and protein secretion were unaffected. When these rats were allowed to recover for 10 weeks, both fertility and the Na and water reabsorption of the cauda were restored. It is proposed that the chlorinated sugar may affect cpididymal sperm metabolism through an effect on the transport function of the epididymis.     

Effect of interleukin 6 on basal and FSH-stimulated secretion of estradiol and progesterone by human granulosa cells in vitro

Aim: To determine the effects of interleukin-6 (IL-6) on the secretion ofestradiol and progesterone by human granulosa cells in vitro. Methods:Granulosa cells were obtained from infertile patients undergoing IVF ETtreatment and cultured with serum-free supplemented HAM's F10 medium.In the absence or presence of FSH, granulosa cells were treated with differ-ent concentrations of gene recombinant human iterlukin-6 (rhIL-6). Themedia were collected after 24, 48, 72 and 96 h and assayed for estradiol andprogesterone. IL-6 and R mRNA was determined by means of RNA slotblot. Results: IL-6 had a significant inhibitory effect on estradiol secre-tion, especially in the presence of FSH. IL-6 inhibited the FSH-stimulatedprogesterone secretion, but not the basal progesterone release. The inhibi-tion shows certain degrees of dose- and time-dependency. Conclusion:IL-6 participates in the regulation of ovarian function through its inhibitoryeffect on FSH-stimulated steroidogenesis by granulosa cells.     

All-trans-retinoic acid aggravates cryoglobulin-associated membranoproliferative glomerulonephritis in mice.

BACKGROUND: Transgenic (tg) mice overexpressing thymic stromal lymphopoietin (TSLP) develop mixed cryoglobulinaemia with renal disease closely resembling human cryoglobulinaemic membranoproliferative glomerulonephritis (MPGN), as well as systemic inflammation involving lung, liver and skin as a result of cryoglobulin deposits. We assessed the effect of all-trans-retinoic acid (ATRA), a powerful anti-inflammatory agent, on this model of cryoglobulinaemic MPGN. METHODS: Groups of male TSLP tg mice and wild-type controls were treated with either ATRA (20 mg/kg) or vehicle 3 times weekly by intraperitoneal injection for 4 or 8 weeks, when mice were then sacrificed. Routine histology and immunohistochemistry for collagen IV, alpha-smooth muscle actin, Mac-2 and Ki67 were performed. Immunoglobulin levels were measured by enzyme-linked immunosorbent assay. RESULTS: ATRA unexpectedly exacerbated renal injury in TSLP tg mice with increased glomerular extracellular matrix, mesangial cell activation, glomerular cell proliferation, glomerular macrophage influx and immune complex deposition. Systemic injuries involving liver and lung, and the amount of circulating cryoglobulins were all worsened by ATRA treatment. Furthermore, ATRA resulted in increased IgG1 and IgM levels, the main components of the cryoglobulins in TSLP tg mice, and a manifestation of an enhanced Th2 immune response. CONCLUSIONS: ATRA is not protective but instead aggravates cryoglobulinaemic MPGN and its systemic manifestations in TSLP tg mice. We speculate these findings may be due to augmented production of pathogenic immunoglobulins and/or an enhanced systemic Th2 response. Although disappointing, our results also suggest caution in the application of retinoid therapy to human disease based on the largely positive animal data reported to date.     

伴侣素CCT6A在结肠直肠癌中的表达及其意义

目的：研究真核生物胞质伴侣素6A（chaperonin containing TCP1 complex,CCT6A）在结肠直肠癌中的表达及临床意义。方法：58例已明确诊断的结肠直肠癌病人入选,分别用Western印迹法和免疫组化方法检测结肠直肠癌细胞系和结肠直肠癌标本中CCT6A的表达,并分析此表达与病人临床病理间的关系。结果：结肠直肠癌组织中CCT6A的表达明显高于结肠直肠的正常组织,具有显著性差异（P〈0．01）。CCT6A的表达与结肠直肠癌浸润深度（P〈0．01）和肿瘤大小（P〈0．05）相关。结论：CCT6A在结肠直肠癌组织中高表达,并与肿瘤浸润深度和肿瘤大小有关,提示CCT6A可作为诊断结肠直肠癌的潜在标记物。     

Hyperhomocysteinaemia in end-stage renal failure patients: Effect of low-dose supplementation with folic acid

SUMMARY: Cardiovascular disease (CVD) is a leading cause of mortality and morbidity in end-stage renal failure (ESRF) patients. In subjects with normal renal function, homocysteinaemia is an independent risk factor for CVD. We studied biochemical determinants of homocysteine (Hcys) levels in 71 ESRF patients on maintenance haemodialysis (HD) treated with recombinant human erythropoietin and supplemented with B-group vitamins. One third of subjects were supplemented with 300–500 μg folic acid daily. A reference range for Hcys was determined in 103 apparently healthy adults without renal impairment. Although blood folate and cobalamin levels were generally in the normal range, most (82%) HD subjects had plasma Hcys levels above the reference range. Significant inverse correlations were noted between plasma Hcys and serum folate ( r =−0.32, P =0.009) and red cell folate ( r =−0.33, P =0.004). Homocysteine concentrations were 23% lower in subjects receiving folk acid supplements (33.9 [SD, 13.6] vs 44.2 [SD, 26.7] μmol/L; P =0.075). No correlations were evident between Hcys and age, time on dialysis, serum cobalamin or measures of vitamin B₆ status (red cell aspartate transaminase or the pyridoxal effect). This study supports the supplementation of HD subjects with folic acid as a measure to reduce plasma Hcys levels and hence possibly atherogenic risk.     

IL-6对人精子顶体反应影响机制的初步探讨

目的:探讨IL-6对人精子顶体反应(AR)的影响机制。方法:采用BAEE/ADH法测定精子顶体酶的活性,以及通过FITC-PSA法检测精子顶体反应。结果:IL-6可诱导精子顶体酶及超氧化物歧化酶(SOD)的活性,促进精子顶体反应;胞外Ca2+单独不能诱导精子顶体反应,且没有胞外Ca2+的参与,IL-6也不能诱导精子顶体反应;蛋白激酶C(PKC)抑制剂calphC能逆转IL-6诱导的精子顶体反应。结论:IL-6对精子顶体反应有一定的促进作用,可能通过诱导精子的顶体酶和SOD活性等途径来实现,在此作用中,也涉及了PKC的激活,且还需要外源性Ca2+的参与。