Influence of Ukrain and cyclophosphamide administration on HA-1 murine hepatoma and LS lymphoma on aspartic proteinase cathepsin D

Cathepsin D, the major lysosomal aspartyl proteinase and a mediator of interferon-gamma and tumor necrosis factor-alpha-induced apoptosis, was studied in murine models of LS lymphosarcoma treated by cyclophosphamide (possible apoptosis induction), and HA-1 hepatoma treated by Ukrain (positive antitumor effect). It was found that cyclophosphamide, as well as cyclophosphamide plus Ukrain, increased cathepsin D specific activity in mice with LS lymphosarcoma. Ukrain alone had no effect on cathepsin D activity in LS lymphosarcoma. In HA-1 hepatoma cells cathepsin D activity was not changed compared with intact normal murine liver (day 10) and activity decreased during tumor development (on day 12). Ukrain significantly increased cathepsin D activity in ascitic fluid (day 10) and had a tendency to increase cathepsin D activity in ascitic cells but not to the normal value.     

Chitotriosidase as a new marker of macrophage stimulation in a tumor model treated with cyclophosphamide and Ukrain

Ukrain has previously been demonstrated to exert a malignotoxic effect in vivo. This antitumor drug has been effective in the treatment of some malignancies in experimental animals as a result of immunostimulation (macrophage stimulation). In the present study, serum chitotriosidase activity was measured as a biochemical marker of macrophage stimulation in several murine and rat models of macrophage stimulation. It was shown that zymosan, carboxymethylated glucan and Triton WR 1339 administration to CBA mice or Wistar rats was followed by a considerable increase in serum chitotriosidase activity. Murine LS lymphosarcoma development decreased serum chitotriosidase activity. Antitumor treatment by Ukrain or cyclophosphamide did not restore this index to the normal value.     

Protein kinase C involvement in apoptosis

• 1.1. Conflicting observations on the involvement of PKC in apoptosis point to a great variability depending on cell type, agent or condition causing apoptosis, phase of the cell cycle and intracellular signaling pathway.
• 2.2. Inhibition by PKC of store-operated calcium entry mechanisms, which are sensitive to the oncoprotein bcl-2, should block the activation of calcium-dependent enzymes triggering the apoptotic cell death.
• 3.3. Activation of phosphatases by ceramide and inhibition of PKC by sphingosine seem to mediate the sphingomyelin pathway to apoptosis.
• 4.4. A putative target protein appears to be p34^cdc2 which is regulated by a network of kinases and phosphatases. The uncoupling of timing for p34^cdc2 activation and the completion of DNA replication results in the so-called “mitotic catastrophe” that shares some features with apoptosis.

     

The influence of Ukrain on the growth of HA-1 tumor in mice: the role of cysteine proteinases as markers of tumor malignancy

The influence of Ukrain (exerting a malignotoxic effect in vivo according to Nowicky J.W. et al [1]) on the growth of Hepatoma A (HA-1) tumors in mice and cysteine proteinases as markers of tumor growth and malignancy was studied. Ukrain administration (0.5 mg per mice of 20 g, in each of 5 i.p. injections) resulted in a reproducible and significant retardation of HA-1 tumor growth in the liver and a prolongation of lifespan compared with the untreated control. Repeated Ukrain administration to mice increased the number of macrophages in ascites, decreased the number of tumor cells, and concomitantly reduced the increased level of monocytes in peripheral blood. In ascitic cells the specific activity of cathepsin B was higher than in the intact liver and was not influenced by Ukrain; activity of cysteine proteinases studied in ascitic fluid was much higher than that in serum. Tumor growth was followed by a decrease in cathepsin B activity in the liver and serum at day 10. Ukrain administration has a tendency to normalize the disturbances of cathepsin B activity during tumor development. The cathepsin L activity changes in ascitic fluid were more impressive than those of cathepsin B, indicating the special role of this cysteine proteinase in HA-1 tumor growth and invasiveness. The role of cysteine proteinases as markers of tumor malignancy and invasiveness is discussed.     

T cell kinetics and apoptosis in immune organs and mammary tumors of rats treated with cyclophosphamide and soluble tumor-associated antigens

The aim of this review was to analyze the role of cell kinetics and apoptosis in the cellular mechanism underlying the effects of soluble tumor-associated antigens (sTAA) on chemically-induced mammary cancer in rats treated with the anticancer drug cyclophosphamide (CPA). Mammary gland cancer was induced with dimethylbenz(a)anthracene (DMBA), and tumors, the spleen, thymus, bone marrow and lymph nodes were studied by morphometric and immunohistochemical methods. We suggest that inhibition of the functional activity of the immune system is one of the main reasons for the toxic effects of CPA, and that the tumor-suppressive antitoxic effects of sTAA result from their activation of T-lymphocyte production in this system, particularly in the spleen and lymph nodes. The results of our experiments have shown that vaccination with sTAA actively promotes generation of the host's antitumor immune response. This is manifested in inhibited proliferative activity of the tumor cells, stimulated production of T lymphocytes and an increased rate of apoptosis among tumor cells.     

舒血宁注射液联合格列齐特治疗2型糖尿病的疗效及对同型半胱氨酸和胱抑素C水平的影响

摘 要 目的：观察舒血宁注射液联合格列齐特治疗2型糖尿病的临床疗效,及对同型半胱氨酸和胱抑素C水平的影响,探讨其作用机制。方法: 2型糖尿病患者60例随机分为观察组和对照组各30例。对照组给予格列齐特降糖治疗,观察组在此基础上给加用舒血宁注射液治疗。疗程均为2周。比较两组疗效、药品不良反应,以及治疗前后患者同型半胱氨酸（Hcy）、胱抑素C（CysC）和血糖（FBG、PBG）水平变化。结果: 观察组总有效率为96.67%,明显高于对照组的63.33%（P＜0.05）。治疗后,两组患者的血糖以及Hcy、CysC水平均较治疗前明显下降（P＜0.05）,且观察组上述指标均低于对照组（P＜0.05）。两组患者药品不良反应发生率比较,差异无统计学意义（P>0.05）。结论:舒血宁注射液对2型糖尿病患者有较好疗效,可以有效降低同型半胱氨酸和胱抑素C水平,更好的控制血糖。     

Induction of apoptosis in cultured Chinese hamster ovary cells by Ukrain and its synergistic action with etoposide

The induction of apoptosis by Ukrain, a novel antitumor drug, was studied in Chinese hamster ovary (CHO) cells bearing multiple copies of recombinant human erythropoietin gene incorporated into their genome (cell lines CHO-k38 and -k38/12). Ukrain was found to be capable of the in vitro induction of apoptosis in the cell lines studied. The effect was less expressed in cells with type I multiple drug resistance (k38/12). Ukrain acted synergistically with etoposide, i.e., the combined effect of both agents was evident at significantly reduced concentrations. This suggests that pharmacological compositions of the drugs may reduce the effective doses used in chemotherapy and thus significantly diminish its toxic side effects. Ukrain was found to exert an unusual effect, manifested as the inhibition of protein secretion by target cells. This phenomenon may be used for the express determination of cell sensitivity to colchicine-like cytostatics, including Ukrain.     

Increased pulmonary collagen synthesis in mice treated with cyclophosphamide

Cyclophosphamide is a widely used anticancer drug which damages lung tissue and elicits a progressive fibrotic lesion in mice. The early time course of collagen accumulation and the changes in collagen synthesis which may accompany this lung damage and fibrosis have not been reported. The total lung concentration of hydroxyproline, an index of collagen content and the extent of pulmonary fibrosis, was not significantly increased in mice until 12 days after treatment with 200 mg/kg cyclophosphamide. The change in total lung hydroxyproline content was preceded, on Day 6, by an increase in the rate of acid-insoluble hydroxyproline synthesis. This rate remained elevated to Day 21, but was no longer significantly increased by Day 28. The percentage of total protein synthesis devoted to the production of collagen was significantly increased 9 and 12 days after cyclophosphamide, compared to saline-treated controls. The rate of pulmonary non-collagen protein synthesis was significantly decreased 3 days after cyclophosphamide, and increased 6 and 15 days after this treatment. These data indicate that cyclophosphamide-induced increases in pulmonary collagen synthesis precede the accumulation of this protein. Increased collagen synthesis may occur in the absence of changes in overall protein synthesis although cyclophosphamide also alters non-collagen protein synthesis.     

瞬时受体电位香草醛亚家族1对高脂处理肝细胞凋亡的保护作用

目的 研究瞬时受体电位香草醛亚家族1(TRPV1)在高脂处理肝细胞中的变化及其作用.方法 将肝细胞(BRL-3A)分为对照组和250、500 μmol·L-1软脂酸钠组,加入辣椒素和辣椒卓平对肝细胞进行干预,通过Q-PCR法测量肝细胞中TRPV1的表达水平,并采用流式细胞仪检测肝细胞的凋亡水平.结果 肝细胞的凋亡比例随软脂酸钠浓度的增加而增加,其浓度为500 μmol·L-时,TRPV1的表达水平明显增加;辣椒素能够上调TRPV1的表达水平并减轻高脂所诱导的肝细胞凋亡;而辣椒卓平能够逆转辣椒素对肝细胞的保护作用.结论 高脂能够激活肝细胞中的TRPV1通道,同时,上调TRPV1,减少高脂诱导的肝细胞凋亡.     

半胱氨酸蛋白酶抑制剂C与儿童肾功能的相关性研究

目的：探讨半胱氨酸蛋白酶抑制剂C（ CysC）与儿童肾功能相关性,并分析CysC与年龄、性别的相关性。方法从LIS系统导出2010-2013年包含CysC的1335名儿童生化筛查组合检测数据,通过排除条件,筛选出901名儿童为参考人群,其中男485名,女416名。 CysC的测定采用液相透射比浊法在Roche DPP全自动生化分析仪上进行。结果901名本地区儿童血清CysC浓度对数呈正态分布,其男女间差异无统计学意义（ P＞0．05）。2岁前血清CysC水平变化较大,其中1个月内CysC水平最高,以后随着年龄增长而逐渐降低,但在2-16岁总体趋于稳定。结论儿童血清CysC参考区间与性别无密切相关性,但不同年龄段之间变化较大,临床应用CysC评价儿童肾功能时应考虑年龄因素。     

Gentamicin-induced apoptosis in LLC-PK1 cells: involvement of lysosomes and mitochondria

Gentamicin accumulates in lysosomes and induces apoptosis in kidney proximal tubules and renal cell lines. Using LLC-PK1 cells, we have examined the concentration- and time-dependency of the effects exerted by gentamicin (1-3 mM; 0-3 days) on (i) lysosomal stability; (ii) activation of mitochondrial pathway; (iii) occurrence of apoptosis (concentrations larger than 3 mM caused extensive necrosis as assessed by the measurement of lactate dehydrogenase release). Within 2 h, gentamicin induced a partial relocalization [from lysosomes to cytosol] of the weak organic base acridine orange. We thereafter observed (a) a loss of mitochondrial membrane potential (as from 10 h, based on spectrophotometric and confocal microscopy using JC1 probe) and (b) the release of cytochrome c from granules to cytosol, and the activation of caspase-9 (as from 12 h; evidenced by Western blot analysis). Increase in caspase-3 activity (assayed with Ac-DEVD-AFC in the presence of z-VAD-fmk]) and appearance of fragmented nuclei (DAPI staining) was then detected as from 16 to 24 h together with nuclear fragmentation. Gentamicin produces a fast (within 4 h) release of calcein from negatively-charged liposomes at pH 5.4, which was slowed down by raising the pH to 7.4, or when phosphatidylinositol was replaced by cardiolipin (to mimic the inner mitochondrial membrane). The present data provide temporal evidence that gentamicin causes apoptosis in LLC-PK1 with successive alteration of the permeability of lysosomes, triggering of the mitochondrial pathway, and activation of caspase-3.     

Protein kinase C involvement in aloe-emodin- and emodin-induced apoptosis in lung carcinoma cell.

1. This study demonstrated aloe-emodin- and emodin-induced apoptosis in lung carcinoma cell lines CH27 (human lung squamous carcinoma cell) and H460 (human lung non-small cell carcinoma cell). Aloe-emodin- and emodin-induced apoptosis was characterized by nuclear morphological changes and DNA fragmentation. 2. During apoptosis, an increase in cytochrome c of cytosolic fraction and activation of caspase-3, identified by the cleavage of its proform, were observed. 3. To elucidate whether the expression of protein kinase C (PKC) isozymes are involved in aloe-emodin- and emodin-induced apoptosis, this study examined the changes of PKC isozymes by Western blotting techniques during aloe-emodin- and emodin-induced apoptosis. 4. The expression of PKC isozymes involved in aloe-emodin- and emodin-induced apoptosis of CH27 and H460 cells. In this study, aloe-emodin and emodin induced the changes of each of PKC isozymes in CH27 and H460 cells. 5. The decrease in the expression of PKC delta and epsilon may play a critical role in aloe-emodin- and emodin-induced apoptosis in CH27 and H460 cells. 6. The present study also demonstrated that PKC stimulation occurs at a site downstream of caspase-3 in the emodin-mediated apoptotic pathway.     

Preliminary results of individual therapy of chronic hepatitis C by Ukrain and interferon-alpha

The effects of Ukrain and recombinant human interferon-alpha 2b (IFN) on the state of the thiol-disulfide ratio (SH/SS) of the blood (Russian Federation patent no. 2150700) were studied in vitro using the amperometric titration method. The blood of 73 chronic hepatitis C (CHC) hepatitis C virus (HCV)-RNA-positive patients was examined. Ukrain was tested in doses of 0.05-2.0 micrograms/ml and IFN in 20-1000 U/ml of blood. After in vitro examination, 59 patients were treated: 28 with Ukrain and 31 with IFN. The first group of 16 patients (including eight with HCV genotype 1b) was treated with individually selected optimal doses of Ukrain (0.5-2.5 mg every second day). The second group of 12 patients was treated with doses of 2.5 mg Ukrain independent of in vitro test results. The third group of 31 patients was treated with individually selected optimal doses of IFN (0.5-2 MU 3 times a week). It was found that 79.4% of CHC patients were sensitive to Ukrain in vitro and 65.1% were sensitive to IFN. CHC patients with genotype 1b were sensitive to IFN only in 16.7% of cases while the figure for Ukrain was 92.3%. CHC patients with other HCV genotypes (3, 1a, 2) were sensitive to Ukrain in 86.7% of cases and to IFN in 70.6%. After 1 month of individual therapy with Ukrain, 87.5% of CHC patients, including six of eight cases with HCV genotype 1b, became PCR-HCV negative. In the group receiving the standard dose of Ukrain, virological response was only 33.3%. After 1 month, 74.2% of CHC patients treated with individual doses of IFN became PCR-HCV negative and after 3 months 90.3% were PCR-HCV negative. The prognostic significance of the method for screening preparations for the treatment of CHC patients was 89.8%. Treatment with Ukrain was without serious negative effects and the number of side effects of IFN in individual therapy was significantly reduced. Ukrain can be used in the treatment of CHC patients, alone or in combination with IFN preparations; in the cases with HCV genotype 1b Ukrain seems more promising than IFN. Individual therapy with Ukrain and IFN increased the efficacy of treatment 2.5-fold in comparison with standard monotherapy with the same preparations, significantly decreased the number of side effects and dramatically improved cost-effectiveness.     

心房颤动患者血清半胱氨酸蛋白酶抑制剂C变化及其与肾素-血管紧张素-醛固酮系统的关系

目的 探讨持续性心房颤动(CAF)患者血清半胱氨酸蛋白酶抑制剂C(CysC)水平的变化及其与肾素-血管紧张素-醛固酮系统(RAAS)的关系.方法 将103例符合入选标准的患者分为CAF组53例和对照组50例.比较2组血清CysC、肾素(PRA)、血管紧张素Ⅱ(Ang Ⅱ)、醛固酮(Ald)水平.结果 与对照组比较,CAF组血清CysC、PRA、AngⅡ、Ald水平均明显升高(P＜0.01);且CAF组血清CysC水平与PRA、Ang Ⅱ、Ald水平呈显著正相关(r分别为0.515、0.489、0.449).结论 CAF患者血清CysC水平明显升高,与其RAAS的激活密切相关.     

Modulation by a human interferon of antitumor effects of cyclophosphamide against a lymphosarcoma in hamsters

Administration to hamsters of a highly purified human leukocyte interferon subtype, IFN-alpha A, obtained by recombinant DNA methods, abolished the efficacy of high doses of cyclophosphamide (2.5 mg/hamster) against the TBD 932 lymphosarcoma. The effect was more pronounced with concomitant than with sequential treatments and did not occur with melphalan. Antagonistic effects occurred at high interferon doses (10(5) I.U./treatment), but an additive or synergistic positive effect occurred at lower interferon doses (10(3) I.U./treatment) and at lower, non-protective, doses of cyclophosphamide. These effects may be due to immunomodulatory responses induced by the drugs involved.     

心房颤动患者血清半胱氨酸蛋白酶抑制剂C变化及其与肾素—血管紧张素—醛固酮系统的关系

目的探讨持续性心房颤动(CAF)患者血清半胱氨酸蛋白酶抑制剂C(CysC)水平的变化及其与肾素—血管紧张素—醛固酮系统(RAAS)的关系。方法将103例符合入选标准的患者分为CAF组53例和对照组50例。比较2组血清CysC、肾素(PRA)、血管紧张素Ⅱ(AngⅡ)、醛固酮(Ald)水平。结果与对照组比较,CAF组血清CysC、PRA、AngⅡ、Ald水平均明显升高(P<0.01);且CAF组血清CysC水平与PRA、AngⅡ、Ald水平呈显著正相关(r分别为0.515、0.489、0.449)。结论 CAF患者血清CysC水平明显升高,与其RAAS的激活密切相关。     

Possible involvement of oxidative stress in cisplatin-induced apoptosis in LLC-PK1 cells

Use of cisplatin, a chemotherapeutic agent, is associated with toxicity as a significant number of patients develop a decline in renal function. The mechanisms by which cisplatin produces renal injury are not well understood. It has been suggested that free radical-catalyzed lipid peroxidation can induce apoptosis or necrosis leading to renal injury. This study examined whether low concentrations of cisplatin induce apoptosis in LLC-PK1 cells and whether caspases 1, 2, 3, 8, and 9 are activated during this event. Our results show a dose- and time-dependent induction of apoptosis by micromolar concentrations of cisplatin. Expression of oncogenes c-myc and p53 was induced, and except for caspase 1, all the other caspases tested were activated. Z-VAD, the broad-spectrum inhibitor of caspases, prevented caspase activation and apoptosis, but not c-myc and p53 induction. On the other hand, N-acetylcysteine prevented cisplatin-induced apoptosis as well as c-myc induction but not p53 induction. The antioxidant trolox also prevented cisplatin-induced apoptosis. The results suggest that antioxidants and caspase inhibitors may alleviate cisplatin-associated nephrotoxicity.     

Antiemetic effectiveness of ondansetron and granisetron in patients with breast cancer treated with cyclophosphamide.

PURPOSE: The antiemetic effectiveness of ondansetron 8 mg i.v, ondansetron 32 mg i.v, and granisetron 10 microg/kg or 1 mg i.v. as prophylaxis in breast cancer patients regimens was studied. METHODS: Data from six U.S. cancer centers were collected retrospectively for 224 patients who received cyclophosphamide-containing therapy between January 1998 and June 2002. Logistic-regression analysis was used to examine the likelihood of chemotherapy-induced nausea and vomiting (CINV) both on an unadjusted basis and controlling for concomitant radiation therapy and dexamethasone use. RESULTS: Seventy-six patients (34%) received ondansetron 32 mg, 68 (30%) received ondansetron 8 mg, and 80 (36%) received granisetron (either 10 microg/kg or 1 mg). Patients receiving ondansetron 8 mg were 2.5 times as likely to have CINV on an adjusted basis as granisetron recipients (p < 0.01). There was no increase in the risk of CINV with ondansetron 32 mg compared with granisetron. Patients treated with ondansetron 8 mg required more rescue antiemetics and more prophylactic antiemetics in subsequent chemotherapy cycles than patients in the other groups. CONCLUSION: In a retrospective multicenter study, granisetron 1 mg or 10 microg/kg and ondansetron 32 mg appeared more effective than ondansetron 8 mg in preventing acute CINV related to cyclophosphamide therapy.     

他克莫司血药浓度与胱抑素C在肾移植受者中的相关性分析

摘 要 目的：回顾性分析肾移植受者术后他克莫司（Tac）的血药浓度与肾功能指标血清胱抑素C（SCysC）以及血清肌酐（SCr）的相关性。方法: 纳入84例服用Tac的肾移植受者,其中男67例,女17例,收集其Tac血药浓度、SCysC和SCr数据;分别按术后时间、血药浓度分组,将浓度与SCysC、SCr进行Pearson相关分析,比较不同时间段及浓度水平下血药浓度分别与SCysC、SCr的相关性。结果: 术后不同时间段内,Tac血药浓度与SCysC、Scr没有显著相关性（P＞0.05）;不同药物浓度水平组,血药浓度与SCysC、Scr亦没有显著相关性。随着术后时间的延长,Tac血药浓度呈逐渐下降的趋势,SCysC、Scr水平先是降低,术后两年又逐渐上升。结论:Tac血药浓度不影响SCysC和Scr等生化指标对于移植肾功能的评价。