Insomnia and multiple health outcomes: umbrella review of meta-analyses of prospective cohort studies.

OBJECTIVES: To evaluate existing evidence of prospective cohort studies on associations between insomnia and multiple health outcomes. STUDY DESIGN: An umbrella review of meta-analyses of prospective cohort studies. METHODS: A systematic search was undertaken in Pubmed, Embase, Cochrane, and Web of Science from inception to October 2021 to find meta-analyses of prospective cohort studies investigating the association of insomnia with any health outcome. The summary relative risk (SRR) for each meta-analysis was recalculated with random-effects model. The methodological quality and the quality of evidence were assessed by the A Measurement Tool to Assess Systematic Reviews and Grading of Recommendations, Assessment, Development and Evaluation, respectively. RESULTS: A total of 25 published meta-analyses of prospective cohort studies, reporting 63 SRRs for 29 unique outcomes were included. Insomnia was mainly related to cardiovascular outcomes and mental disorders. The former comprised atrial fibrillation (SRR: 1.30, 95% confidence interval: 1.26 to 1.35), cardiovascular diseases (1.45, 1.29 to 1.64), coronary heart disease (1.28, 1.10 to 1.50), myocardial infarction (1.42, 1.17 to 1.72), and stroke (1.55, 1.39 to 1.72). The latter involved alcohol abuse (1.35, 1.08 to 1.67), all mental disorders (2.16, 1.70 to 3.97), anxiety (3.23, 1.52 to 6.85), depression (2.31, 1.90 to 2.81), suicidal ideation (2.26, 1.79 to 2.86), suicidal attempt (1.99, 1.31 to 3.02), and suicidal death (1.72, 1.42 to 2.08). Besides, insomnia enhanced the risk of Alzheimer's disease (1.51, 1.06 to 2.14) and hyperlipidemia (1.64, 1.53 to 1.76). CONCLUSION: Insomnia exhibits considerable adverse outcomes, primarily comprises cardiovascular outcomes and mental disorders, but further studies with robustly designed trials are needed to draw firmer conclusions.

[Detection of circulating tumor cells with chromosomes 7 and 8 polysomy in non-small cell lung cancer and its correlation with epidermal growth factor receptor mutations in cancer tissue].

Objective: To investigate the value of chromosomes 7 and 8 polysomy in circulating tumor cells (CTCs) for the diagnosis of non-small cell lung cancer, and the correlation of CTCs with clinical pathological characteristics and epidermal growth factor receptor (EGFR) mutations in cancer tissue. Methods: Fifty-seven patients with non-small cell lung cancer and 21 patients with benign lung diseases were enrolled at Beijing Chaoyang Hospital, Capital Medical University, Beijing, China from November 2017 to October 2020. Negative enrichment combined with immunofluorescence in situ hybridization (imFISH) was used to identify CTCs polysomy on chromosomes 7 and 8. EGFR mutations in 56 lung cancer patients was detected using ARMS-PCR. Results: CTCs were detected in 93.0% (53/57) of non-small cell lung cancers and 28.6% (6/21) benign lung lesions. The difference between lung cancer patients and the control cohort was statistically significant (P<0.01). Receive operator curve (ROC) analyses showed that, when the cut-off value was 1 cell/3.2 mL, Youden index had the highest sensitivity of 93.0% and specificity of 71.4% (AUC=0.906, 95%CI:0.833-0.980, P<0.01). The positive rate of CTCs in stage Ⅲ-Ⅳ cancers was significantly higher than that in stage Ⅰ-Ⅱ (P=0.023). No significant correlation was observed between positive rate of CTCs or chromosome polysomy and age, gender, smoking status, pathologic types and EGFR mutation status. The number of CTCs in EGFR mutated group was higher than that in the non-mutated group (6.5±1.1 vs. 3.7±0.7, P=0.045). The detection rate for CTCs ≥5 in the EGFR mutated group was also higher than the EGFR non-mutated group (52.0% vs. 19.4%,P=0.010). Conclusion: Detection of CTCs with chromosomes 7 and 8 polysomy has potential value in auxiliary diagnosis of non-small cell lung cancer, and the number of CTCs is correlated to TNM stage and EGFR gene mutation status.

[Clinical auxiliary diagnosis value of high frequency ultrasonographic measurements of the thickness of transverse carpal ligaments in carpal tunnel syndrome patients].

OBJECTIVE: To evaluate the meaning and value of high-frequency ultrasound in the diagnosis of carpal tunnel syndrome (CTS). METHODS: In this study, 48 patients (unilateral hand) with CTS were analyzed. The thickness of transverse carpal ligaments at the pisiform bone was measured using high-frequency ultrasound. Open carpal tunnel release procedure was performed in the 48 CTS patients, and the thickness of transverse carpal ligaments at the hamate hook bone measured using vernier caliper under direct vision. The accuracy of thickness of transverse carpal ligaments was evaluated using high-frequency ultrasound. high-frequency ultrasound measurement of thickness of transverse carpal ligaments at the hamate hook bone and pisiform bone, and determination of the diagnostic threshold measurement index using receiver operating characteristic (ROC) curve, sensitivity and specificity were performed and the correlation between the thickness of transverse carpal ligaments and nerve conduction study (NCS) analyzed. RESULTS: The thickness of transverse carpal ligaments in the CTS patients were (0.42±0.08) cm (high-frequency ultrasound) and (0.41±0.06) cm (operation) at hamate hook bone, and there was no significant difference between the two ways (t=0.672, P>0.05). The optimal cut-off value of the transverse carpal ligaments at hamate hook bone was 0.385 cm, the sensitivity 0.775, and the specificity 0.788. The optimal cut-off value of the transverse carpal ligaments at the pisiform bone was 0.315 cm, the sensitivity 0.950, and the specificity 1.000. The transverse carpal ligaments thickness and wrist-index finger sensory nerve conduction velocity (SCV), wrist-middle finger SCV showed a negative correlation. CONCLUSION: High frequency ultrasound measurements of thickness of transverse carpal ligaments is a valuable method for the diagnosis of CTS.

Telomerase expression in human somatic cells does not induce changes associated with a transformed phenotype.

Expression of the human telomerase catalytic component, hTERT, in normal human somatic cells can reconstitute telomerase activity and extend their replicative lifespan. We report here that at twice the normal number of population doublings, telomerase-expressing human skin fibroblasts (BJ-hTERT) and retinal pigment epithelial cells (RPE-hTERT) retain normal growth control in response to serum deprivation, high cell density, G1 or G2 phase blockers and spindle inhibitors. In addition, we observed no cell growth in soft agar and detected no tumour formation in vivo. Thus, we find that telomerase expression in normal cells does not appear to induce changes associated with a malignant phenotype.

Effect of dietary protein and energy intake on embryonic survival and gene expression in the uterine endometrium of early pregnant gilts.

Porcine embryonic loss during early gestation is a serious problem in swine production. Improving embryonic survival can be achieved by maternal manipulation. Protein and energy are two major components of the diet, which play decisive roles in embryonic survival. This study was performed to evaluate the effects of enhancing maternal protein or energy intake on embryonic survival during early gestation in gilts and to explore the underlying mechanism. From day (d) 0 to 30 of gestation, 40 gilts (Landrace × York) were randomly allocated to 5 diets according to daily intake of low (L, National Research Council (NRC) recommendation for gestation gilts), medium (M, 20% higher than NRC) or high (H, 40% higher than NRC) CP or metabolisable energy (ME) (LCPLME, MCPLME, HCPLME, LCPHME, HCPHME). Gilts were sacrificed on d 30 of gestation, and number of foetuses and corpora lutea, embryonic survival rate, uterine weight, and total volume of allantoic fluid were recorded or calculated. Gene expression was determined by Quantitative Real-time PCR (qPCR), western blot or immunohistochemistry. Results showed that increasing protein or ME intake significantly increased embryonic survival rate. Compared with diet LCPLME, plasma progesterone (P4) concentration in diet LCPHME increased at d 14 and d 30 of gestation. Progesterone receptor (PGR) was found not to be expressed in the epithelia but was strongly expressed in the stroma of the endometrium. Increasing protein or ME intake did not alter PGR expression in the endometrium. There was also no change in the amount of P4, hepatocyte growth factor, and fibroblast growth factor-7 in the endometrium. The mRNA abundance of cationic amino acid transporter 1 in the endometrium in diet LCPHME and HCPHME was significantly lower than in diet LCPLME. Diet HCPLME showed a tendency to increase neutral amino acid transporter 1 mRNA expression in the endometrium compared to diet LCPLME (P = 0.087). In conclusion, increasing maternal protein or ME intake had a positive effect on the embryonic survival. Increased protein intake by 20 or 40% did not alter plasma P4 level, but increasing ME intake by 40% improved plasma P4 concentration at d 14 and 30 of gestation. Increasing maternal protein or ME intake did not induce PGR expression in the endometrium. Maternal protein and energy intake likely mediate transportation of cationic and neutral amino acids from mother to foetus to affect embryonic survival and development.

Effects of soybean isoflavones on the growth performance, intestinal morphology and antioxidative properties in pigs.

Soybean meal is rich in soybean isoflavones, which exhibit antioxidant, anti-inflammatory, antiviral and anticancer functions in humans and animals. This study was conducted to investigate the effects of soybean isoflavones on the growth performance, intestinal morphology and antioxidative properties in pigs. A total of 72 weaned piglets (7.45 ± 0.13 kg; 36 males and 36 females) were allocated into three treatments and fed corn-soybean meal (C-SBM), corn-soy protein concentrate (C-SPC) or C-SPC supplemented with equal levels of the isoflavones found in the C-SBM diet (C-SPC + ISF) for a 72-day trial. Each treatment had six replicates and four piglets per replicate, half male and half female. On day 42, one male pig from each replicate was selected and euthanized to collect intestinal samples. The results showed that compared to pigs fed the C-SPC diet, pigs fed the C-SBM and C-SPC + ISF diets had higher BW on day 72 (P < 0.05); pigs fed the C-SBM diet had significantly higher average daily gain (ADG) during days 14 to 28 (P < 0.05), with C-SPC + ISF being intermediate; pigs fed the C-SBM diet tended to have higher ADG during days 42 to 72 (P = 0.063), while pigs fed the C-SPC + ISF diet had significantly higher ADG during days 42 to 72 (P < 0.05). Moreover, compared to pigs fed the C-SPC diet, pigs fed the C-SBM diet tended to have greater villus height (P = 0.092), while pigs fed the C-SPC + ISF diet had significantly greater villus height (P < 0.05); pigs fed the C-SBM and C-SPC + ISF diets had significantly increased villus height-to-crypt depth ratio (P < 0.05). Compared with the C-SPC diet, dietary C-SPC + ISF tended to increase plasma superoxide dismutase activity on days 28 (P = 0.085) and 42 (P = 0.075) and reduce plasma malondialdehyde (MDA) content on day 42 (P = 0.089), as well as significantly decreased jejunal mucosa MDA content on day 42 (P < 0.05). However, no significant difference in the expression of tight junction genes among the three groups was found (P > 0.05). In conclusion, our results suggest that a long-term exposure to soybean isoflavones enhances the growth performance, protects the intestinal morphology and improves the antioxidative properties in pigs.

Long non-coding RNA HULC promotes proliferation and osteogenic differentiation of bone mesenchymal stem cells via down-regulation of miR-195.

OBJECTIVE: LncRNAs HULC has been reported to be important regulators in the development of various human diseases. However, the role of HULC in bone mesenchymal stem cells (BMSCs) remains unclear. The present study aimed to explore the regulatory effect of HULC on proliferation and osteogenic differentiation of BMSCs and the underlying mechanism. MATERIALS AND METHODS: The expression of HULC and miR-195 in BMSCs were altered by transfection and measured by qRT-PCR. Cell viability was measured by the CCK-8 assay. Osteogenic differentiation of BMSCs was determined by evaluation of osteogenic markers (Ocn, ALP, Runx2, and Col-1) expression levels using Western blot and qRT-PCR. Furthermore, Western blot was performed to assess the expression of proliferation-related factors, Wnt/ß-catenin and p38MAPK pathway-related factors. RESULTS: HULC overexpression significantly increased cell viability, down-regulated p21 expression but up-regulated CyclinD1 expression, and promoted the levels of osteogenic markers. However, the complete opposite effect was observed in HULC knockdown. Notably, miR-195 expression was negatively regulated by HULC and miR-195 exerted a reversed effect of HULC on BMSCs. Moreover, miR-195 mediated the regulatory effect of HULC on BMSCs proliferation and osteogenic differentiation, as miR-195 mimic abolished the effect of HULC overexpression on BMSCs. We also found that HULC overexpression enhanced the activation of Wnt/ß-catenin and p38MAPK pathway through down-regulating miR-195. CONCLUSIONS: We revealed that HULC promoted proliferation and osteogenic differentiation of BMSCs. The potential mechanism might be involved in its negative regulation on miR-195 and enhanced activation of Wnt/ß-catenin and p38MAPK pathway.

Effects of dietary grape proanthocyanidins on the growth performance, jejunum morphology and plasma biochemical indices of broiler chicks.

Grape proanthocyanidins (GPCs) are a family of naturally derived polyphenols that have aroused interest in the poultry industry due to their versatile role in animal health. This study was conducted to investigate the potential benefits and appropriate dosages of GPCs on growth performance, jejunum morphology, plasma antioxidant capacity and the biochemical indices of broiler chicks. A total of 280 newly hatched male Cobb 500 broiler chicks were randomly allocated into four treatments of seven replicates each, and were fed a wheat-soybean meal-type diet with or without (control group), 7.5, 15 or 30 mg/kg of GPCs. Results show that dietary GPCs decrease the feed conversion ratio and average daily gain from day 21 to day 42, increase breast muscle yield by day 42 and improve jejunum morphology between day 21 and day 42. Chicks fed 7.5 and 15 mg/kg of GPCs show increased breast muscle yield and exhibit improved jejunum morphologies than birds in the control group. Dietary GPCs fed at a level of 15 mg/kg markedly increased total superoxide dismutase (T-SOD) activity between day 21 and day 42, whereas a supplement of GPCs at 7.5 mg/kg significantly increased T-SOD activity and decreased lipid peroxidation malondialdehyde content by day 42. A supplement of 30 mg/kg of GPCs has no effect on antioxidant status but adversely affects the blood biochemical indices, as evidenced by increased creatinine content, increased alkaline phosphatase by day 21 and increased alanine aminotransferase by day 42 in plasma. GPC levels caused quadratic effect on growth, jejunum morphology and plasma antioxidant capacity. The predicted optimal GPC levels for best plasma antioxidant capacity at 42 days was 13 to 15 mg/kg, for best feed efficiency during grower phase was 16 mg/kg, for best jejunum morphology at 42 days was 17 mg/kg. In conclusion, GPCs (fed at a level of 13 to 17 mg/kg) have the potential to be a promising feed additive for broiler chicks.

1,25-Dihydroxyvitamin D3 protects human leukemic cells from tumor necrosis factor-induced apoptosis via inactivation of cytosolic phospholipase A2.

The mechanism by which tumor necrosis factor (TNF) induces death of cancer cells appears to involve the activation of cytosolic phospholipase A2 (cPLA2). U937 human leukemic cells treated with 1,25-dihydroxyvitamin D3 [1,25(OH)2D3; 10(-8) M] become resistant to TNF, an effect that is independent of cell cycle status and expression of TNF receptors or BCL-2. In this study, TNF produced a dose- and time-dependent enhancement of [3H]arachidonic acid release in U937 cells. The amount of [3H]arachidonic acid release was positively associated with TNF-induced apoptosis. Both immunofluorescence microscopy and Western blotting of cell subcompartments demonstrated translocation of cPLA2 from the cytosol to the cell membrane in response to TNF. In addition, TNF up-regulated expression of cPLA2 mRNA. An antisense oligonucleotide to cPLA2 and the cPLA2 inhibitor 4-bromophenacyl bromide significantly inhibited TNF-induced cytotoxicity. Prior incubation of cells with 1,25(OH)2D3 significantly inhibited (a) TNF-induced [3H]arachidonic acid release and apoptosis, (b) TNF-induced translocation of cPLA2 to the membrane, and (c) the up-regulation of cPLA2 mRNA with TNF. Furthermore, the inhibitory effect of 1,25(OH)2D3 was not reversed by inhibitors of transcription or translation. The data suggest that activation of cPLA2 is involved in TNF-induced apoptosis of leukemic cells. 1,25(OH)2D3 directly inhibits cPLA2 translocation and mRNA up-regulation induced by TNF. Disruption of cPLA2 activation may represent a possible mechanism whereby leukemic cells can become resistant to TNF-mediated killing.

The effects of a novel synthetic emulsifier product on growth performance of chickens for fattening and weaned piglets.

Two experiments were conducted to evaluate the effects of a novel synthetic emulsifier product (AVI-MUL TOP) on the growth performance of chickens for fattening and weaned piglets. The emulsifier product consists of 50% vegetal bi-distillated oleic acid emulsified with 50% glyceryl polyethyleneglycol ricinoleate. In experiment 1, 480 1-day-old female Cobb500 chickens for fattening were assigned to two treatments: (1) a control diet (CTR); and (2) the control diet+the emulsifier (AMT, 1 g/kg from day 0 to day 10, 0.75 g/kg from day 10 to day 20 and 0.5 g/kg from day 20 to day 34 of the trial). AMT supplementation increased BW on days 20 and 34 (P<0.01). Dietary AMT increased the average daily gain and average daily feed intake (ADFI) from day 10 to day 20, from day 20 to day 34 and from day 0 to day 34 (P<0.01). A reduced feed conversion ratio was observed in the AMT group from day 10 to day 20 (P<0.01). In experiment 2, 96 Stambo HBI×Dalland piglets were weaned at 24 days and assigned to two treatments (the basal diet without the product (CTR) or with 2 g/kg emulsifier from day 0 to day 14 and 1.5 g/kg from day 14 to day 42 (AMT)). There was an increase in the ADFI associated with AMT supplementation from day 14 to day 42 (P=0.04). These results indicated that supplementation with the synthetic emulsifier may significantly improve the growth performance of chickens for fattening and numerically improve that of weaned piglets.

Effect of a post-weaning diet supplemented with functional feed additives on ileal transcriptome activity and serum cytokines in piglets challenged with lipopolysaccharide.

This study evaluated the potential of a weanling diet supplemented with trace minerals, vitamins, prebiotics, essential oils, antioxidants and bovine colostrum (BC) to modulate the inflammatory response of low-weight (LW) and high-weight (HW) piglets challenged with lipopolysaccharide (LPS). At weaning (20±1 d), litters from 32 sows were assigned to four groups: control diet (CTL), CTL plus dietary supplements (DS) or the antibiotic chlortetracycline (ATB), or DS plus BC in place of plasma proteins in the weanling diet (DS+BC). At 37 d (T0), two LW and two HW piglets were bled to evaluate ex vivo cytokine production by LPS activated peripheral blood mononuclear cells (PBMCs). In parallel, LW and HW piglets received intraperitoneal LPS and were bled at slaughter at 4h (T4) or 18h (T18) post-injection. Ileal tissues from these piglets and two unchallenged medium weight (MW) piglets per treatment were excised and analyzed by microarray. At T0, cytokine production of LPS-activated PBMCs was not affected by dietary treatments. At T4 after LPS challenge, serum concentrations of TNF-α, IL-6, IL-8, and IL-10 were increased in all piglets (P<0.01). Interestingly, the LW piglets had a higher TNF-α level than the HW piglets did (P=0.05). Dietary treatments had no effect on the piglet serum concentration of these cytokines neither at T4 nor at T18. Microarray data and QPCR analysis reveal that several genes were differentially expressed in the LPS-challenged piglets in comparison with the two control MW piglets (P<0.001). However, the dietary treatments had a slight effect on the ileal gene expression of the T4 and T18 LPS-challenged piglets when all piglets were included in the analysis. But when body weight (LW and HW) was considered as a fixed effect, the microarray analysis showed that the expression of 54 genes was differentially modulated by the dietary treatments in the T4 and T18 LPS-challenged LW piglets (P<0.05) while in HW piglets no difference was observed. QPCR analyses confirm that the level expression of several genes was reduced in LW piglets fed DS or DS+BC diet compared with ATB piglets. In conclusion, LPS challenge induced a transitional inflammation in weanling piglets that was characterized by increased blood-circulating cytokines and gut transcriptome activity. Results also suggest that the weanling diet supplemented with feed additives attenuated the ileal gene response to the LPS challenge, an effect that was more pronounced in the LW piglets.

Effects of a blend of essential oils and an enzyme combination on nutrient digestibility, ileum histology and expression of inflammatory mediators in weaned piglets.

The aim of this study was to test the hypothesis of an improved gut environment of post-weaning piglets when administered a blend of essential oils (EO; thymol and cinnamaldehyde) and an enzyme combination (xylanase and ß-glucanase (XB)) either alone or in combination. To assess the effect of dietary treatments, faecal nutrient digestibility and microbial counts, as well as ileum histology and gene expression of inflammatory mediators were evaluated. One hundred and ninety-two weaned piglets were allocated into four experimental treatments, and fed the basal diet (CTRL) either without or with EO, XB or their combination (EO+XB) for a 42-day period. The experiment concerning digestibility was designed with two periods (period I: days 15 to 21; period II: days 29 to 35) and the faeces were collected on days 20, 21, 34 and 35. On day 42, six piglets from each treatment were slaughtered. It was found that EO, XB and EO+XB supplementation did not affect (P>0.05) the growth performance of the piglets from days 0 to 42. Moreover, no dietary effect on faecal score was observed. Faecal digestibility of dry matter, organic matter, ash, dietary fibre, lipid, CP and NDF were increased from period I to period II (P<0.01 to P=0.06), while no effect (P>0.05) of EO, XB or their combination on the faecal digestibility was observed at both periods. Compared with the CTRL diet, dietary XB reduced the faecal Lactobacillus and Escherichia coli counts but increased the Lactobacillus to Coliforms ratio on day 42 (P=0.02, 0.03 and 0.03, respectively), and all the additives supplementations decreased the counts of faecal Coliforms on day 42 (P<0.01). XB supplementation increased the villus to crypt ratio (P=0.04) and reduced the mucosal macrophages number (P<0.01) in the ileum compared with the CTRL group, and dietary EO or EO+XB decreased the number of lymphatic follicles (P=0.01 and P<0.01, respectively) and mucosal macrophages (P=0.02 and P<0.01, respectively). In addition, the interleukin (IL)-1α was downregulated in piglets treated with EO+XB compared with the EO group (P=0.02). In conclusion, the administration of EO, XB or their combination was effective in improving ileum histology, and EO+XB supplementation might benefit the modulation of the expression of ileum inflammatory cytokines in piglets.

SL4, a chalcone-based compound, induces apoptosis in human cancer cells by activation of the ROS/MAPK signalling pathway.

OBJECTIVES: SL4, a chalcone-based compound, exhibits clearly inhibitory effects on HIF-1 and has been shown to effectively suppress tumour invasion and angiogenesis in vitro and in vivo. Here, studies were conducted to determine SL4's anti-apoptotic effects and its underlying mechanisms, in human cancer cells. MATERIALS AND METHODS: Cytotoxicity, apoptotic induction and its involved mechanisms of SL4 were investigated using normal cells, cancer cells and mouse xenograft models. The role of reactive oxygen species (ROS) and mitogen-activated protein kinase (MAPK) signalling in SL4-induced apoptosis was explored by manipulating specific scavenger or signalling inhibitors, in cultured cells. RESULTS: SL4 significantly inhibited cell population growth of human cancer cell lines but exhibited lower cytotoxicity against normal cells. In addition, SL4 effectively induced apoptosis of Hep3B and MDA-MB-435 cells by activating procaspase-8, -9 and -3, and down-regulating expression levels of XIAP, but did not affect HIF-1 apoptosis-related targets, Survivin and Bcl-XL. Further study showed that SL4 also reduced mitochondrial membrane potential and promoted generation of ROS. ROS generation and apoptotic induction by SL4 were blocked by NAC, a scavenger of ROS, suggesting SL4-induced apoptosis via ROS accumulation. We also found that MAPKs, JNK and p38, but not ERK1/2, to be critical mediators in SL4-induced apoptosis. SP600125 and SB203580, specific inhibitors of JNK kinase and p38 kinase, significantly retarded apoptosis induced by SL4. Moreover, anti-oxidant NAC blocked activation of JNK and p38 induced by SL4, indicating that ROS may act as upstream signalling of JNK and p38 activation. It is noteworthy that animal studies revealed dramatic reduction (49%) in tumour volume after 11 days SL4 treatment. CONCLUSIONS: These data demonstrate that SL4 induced apoptosis in human cancer cells through activation of the ROS/MAPK signalling pathway, suggesting that it may be a novel lead compound, as a cancer drug candidate, with polypharmacological characteristics.

TNF-mediated killing of human leukaemic cells: effects of endogenous antioxidant levels and TNF alpha expression in leukaemic cell lines.

Using the human erythroleukaemic cell line K562 cl.6 and its daunorubicin-resistant subline K/DAU600, and the human T-lymphoblastic leukaemic cell line CCRF-CEM and its vinblastine-resistant subline CEM/VLB100, we have shown that the drug-resistant cell lines were more sensitive to cytotoxicity induced by tumour necrosis factor-alpha (TNF alpha). Drug-resistant cell lines showed increased activities of copper/zinc superoxide dismutase (Cu/ZnSOD) and catalase compared with their parental drug-sensitive cell lines. However, the greater susceptibility of drug-resistant cells to TNF alpha cytotoxicity was, in part, related to their decreased activities of manganese superoxide dismutase (MnSOD). Persistence of this differential sensitivity when MnSOD is inhibited by sodium nitroprusside (SNP) suggests that the greater susceptibility of drug-resistant cells to TNF alpha was not entirely due to their decreased level of MnSOD activity. K562 cl.6 and K/DAU600, which were more resistant to TNF alpha, both expressed greater levels of endogenous plasma membrane-bound TNF alpha than the CCRF-CEM cell line. All cell lines examined were (more or less) equal in susceptibility to the cytolytic effect of exogenous O2-. generated by xanthine/xanthine oxidase. These results demonstrate that both MnSOD and endogenous TNF alpha play a role in protecting leukaemic cells against TNF alpha cytotoxicity, but there is an unknown mechanism that causes drug-resistant cells to be more susceptible to TNF alpha cytotoxicity.

The anti-leukaemic effects and the mechanism of sodium selenite.

The anti-proliferative effects of selenium were studied both in vivo and in vitro. At a selenium concentration of 0.6 micrograms/ml, cells from patients with ALL-L1, L2 and AML-M1, M3 and M5 were more sensitive than cells from patients with CML. Cells from patients with AML-M2, CLL and leukaemic lymphoma were least sensitive. Normal bone marrow or peripheral blood cells were not sensitive to selenium at this concentration. In the mouse leukaemia models (L797, L615, L7712), the sensitivity of leukaemic cells were: L797 (93% cytotoxicity) greater than L615 (49.7% cytotoxicity) greater than L7712 (4.4% cytotoxicity). Sodium selenite injected i.p. increased the longevity of L797-inoculated mice. Administration of 40 micrograms selenium daily for 7 days resulted in a significant increase in the longevity of mice inoculated with 10(5) L797 cells. However, no remarkable increase of the longevity was observed in either L615- or L7712-inoculated mice after treatment with sodium selenite for 7 days. Treatment of the HL-60 leukaemic cell line with selenium caused a dose- and time-related decrease in DNA, RNA and protein syntheses as measured by [3H]-thymidine, [3H]-uridine and [3H]-leucine uptake respectively. The inhibitory effect of selenium on DNA synthesis was reversed when selenium was removed from the medium, demonstrating that selenium-induced inhibition of DNA synthesis was due to interference with DNA biosynthesis rather than DNA template damage. These results suggest that the anti-leukaemic effect of sodium selenite is associated with inhibition of DNA replication, transcription and translation.

Modulation of surface TNF expression by human leukaemic cells alters their sensitivity to exogenous TNF.

In this study, U937 leukaemic cells underwent apoptotic cell death following exposure to TNF. Pre-incubation of cells for 48 h with VitD(3) (10(-8)M) induced resistance to TNF, whereas incubation with tau-IFN or GM-CSF increased susceptibility to TNF. Resistance to exogenous TNF (exTNF) following culture with VitD(3) was associated with increased expression of endogenous TNF (enTNF). The TNF inhibitors pentoxifylline(PTF) and dichloroisocoumarin (DCI) inhibited TNF synthesis by U937 cells and abrogated the increase in resistance to TNF seen with VitD(3). The tau-IFN increased TNF expression, whereas GM-CSF had little effect. The data show that the sensitivity of leukaemic cells to exTNF can be modulated by cytokines. The protective effect of VitD(3) is mediated in part by directly upregulating enTNF synthesis.

Evaluation of the in vitro antiproliferative properties of four novel anthracyclines YM1, 3, 4 and 6 in human leukemia cell lines.

The antitumor activity of novel doxorubicin analogues YM1, YM3, YM4 and YM6 was evaluated against drug sensitive U937 monocytic leukemia and CCRF-CEM lymphoid leukemia cell lines, as well as drug resistant CEM/VLB100 lymphoid multidrug resistant leukemia cell line by a [3H]thymidine incorporation assay. Different antileukemic activities of these new anthracyclines were observed in our studies. These novel anthracyclines produced a dose- and time-dependent inhibition in all the leukemic cell lines tested, while YM1 and YM3 were more effective than YM4 and YM6 against all the leukemic cell lines. The antitumor activity of all these novel analogues was lower than that of doxorubicin or epidoxorubicin in drug sensitive leukemic cells. The relative resistance values (IC50 of resistant cell line/IC50 of sensitive parental cell line) of YM1, 3, 4 and 6 were 27, 7, 5 and 14 respectively. These were lower than the resistance values for ADM and EDR which were 45 and 40 respectively. YM3 had a similar antileukemic activity against the CEM/VLB100 drug resistant leukemic cell line to ADM or EDR with a lower relative resistance value and a slightly increased IC50 value. Our results suggest that YM3 may be used in high dose for the clinical treatment of leukemias with possible less cardiotoxicity as well as less drug resistance.

Structure-dependent antitumor activities of novel anthracyclines YM1, YM3, YM4 and YM6: drug transport properties and effects on biomacromolecule synthesis in drug sensitive and resistant leukemia cells.

The antitumor activities of four novel doxorubicin (DOX) analogues, YM1, YM3, YM4 and YM6 in relation to their structure and drug transport properties, have been investigated in U937 monocytic and CCRF-CEM lymphoid drug sensitive leukemia cell lines, as well as in CEM/VLB100, a drug resistant subline displaying high levels of P-glycoprotein. Treatment of all cell lines with YM1, 3, 4 and 6 produced a dose-dependent decrease in DNA, RNA and protein synthesis as measured by [3H]-thymidine, [3H]-uridine and [3H]-leucine uptake respectively. YM1 was more effective than YM3, YM4 or YM6 against the drug sensitive cells. The antitumor effects of all these DOX-analogues on macromolecule synthesis in U937 and CCRF-CEM cells were lower than that of DOX and epirubicin (EDR). A rapid accumulation of the novel anthracyclines was found in all cell lines compared with DOX or EDR. However, the maximal accumulation of the DOX-analogues was lower than that of EDR. There is a greater efflux from CCRF-CEM sensitive cells and less from CEM/VLB100 resistant cells of the DOX-derivatives when compared with EDR and DOX. Drug-induced cytotoxicity significantly correlated (P < 0.05) with drug retention levels in CCRF-CEM and U937 drug sensitive cells as indicated by an inverse correlation curve between anthracycline retention and drug-induced IC50 value. It was demonstrated that an increased level of drug retained within the sensitive cells would therefore produce a more cytotoxic effect of the drug. However, no such correlation was observed in CEM/VLB100 resistant cells. YM3 was shown to have an increased antitumor activity against CEM/VLB100 resistant cells compared with DOX with a lower resistance factor. These results showed that the antitumor effects of four novel DOX-analogues, like DOX or EDR, were associated with inhibition of DNA replication, transcription and translation. The finding that resistant leukemic cells are more susceptible to the cytotoxic effect of YM3 than DOX warrants further investigation to identify the intrinsic mechanism of resistance.