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BACKGROUND Type A insulin resistance syndrome is a rare disorder caused by mutations in the gene encoding the insulin receptor. Its coexistence with ovarian serous papillary cystadenofibroma is even rarer. CASE SUMMARY A 14-year-old girl developed type A insulin resistance syndrome and showed high fasting insulin, glucose, and hemoglobin A1c (HbA1c) levels. The girl suffered from ovarian serous papillary cystadenofibroma. The laboratory results were as follows: fasting insulin was 2624.90 pmol/L and HbA1c was 8.5%. A heterozygous missense mutation on exon 20 of the insulin receptor gene (c.3601C>T, Arg1201Trp) was observed. The histopathological diagnosis was a cystic lesion that extended to the upper right uterus, indicating a right ovarian serous papillary cystadefibroma accompanied by focal interstitial hyperplasia. The patient was treated with metformin for over 6 mo. Additionally, laparoscopic resection (bilateral) of the ovarian lesion and laparoscopic intestinal adhesiolysis were performed under general anesthesia. Diet therapy combined with exercise was then initiated. The patient had an uneventful recovery. The patient also showed improved blood glucose control, with reduced levels of fasting insulin (857.84 pmol/L) and HbA1c (7.0%). CONCLUSION Insulin resistance may play a significant role in the induction of tumors. It is important to investigate further the association between insulin resistance and tumors and the underlying mechanism.

Nonalcoholic fatty liver disease is likely to be associated with increased circulating branched-chain amino acids. We investigated the relationship between changes in branched-chain amino acids levels in the serum and improvement in liver fat content caused by exercise intervention in individuals with nonalcoholic fatty liver disease. The exploratory study included 208 central obesity and nonalcoholic fatty liver disease individuals from an exercise intervention randomized clinical trial for nonalcoholic fatty liver disease. The participants were randomly assigned to control, moderate, and vigorous-moderate exercise groups for 12 months. Changes in total branched-chain amino acids, leucine, isoleucine, and valine levels from baseline to 6 months were calculated. Liver fat content was determined by proton magnetic resonance spectroscopy. Reductions in circulating levels of total branched-chain amino acids, leucine, and valine levels from baseline to 6 months were significantly associated with the improvement of liver fat content at 6 months and 12 months (p p

Objective Hepatitis B virus infection is a major social and economic burden in developing countries, especially in China. We aimed to evaluate the effects of hepatitis B surface antigen (HBsAg) positive status on the pregnancy outcomes in the Chinese population. Methods This retrospective cohort study was performed using data from the Medical Birth Registry of Xiamen, China, from January 2011 to March 2018. Multivariate logistic regression analysis was used to assess the association between the HBsAg status and pregnancy outcomes. Results This study included 3,789 HBsAg-positive women and 29, 648 non-exposed women. The HBsAg-positive pregnant women were slightly older in age (29.3±4.3 vs. 28.9±4.4, P< 0.001). Additionally, pregnant women with a positive HBsAg status had higher odds of gestational diabetes mellitus (GDM) (adjusted odds ratio [aOR], 1.13; 95% confidence interval [CI], 1.03–1.23), and cesarean delivery (aOR, 1.12; 95%CI, 1.03–1.21). The risk of infants being large or small-for-gestational age, having low-birth weight, and of macrosomia, preterm birth, and stillbirth did not differ significantly between the HBsAg-positive and–negative women. Conclusion In Xiamen, China, the slightly higher risk of GDM and cesarean section in women positive for HBsAg should not be neglected. Further studies should be conducted to evaluate the effects of HBsAg positivity on the pregnancy outcomes in different ethnic populations.

The growth trajectory of Chinese preschoolers still remains unclear. Our objective was to determine whether there was an association between adverse pregnancy outcomes and overweight offspring. We analyzed population-based retrospective cohort data from the Medical Birth Registry of Xiamen, which comprised 33,157 children examined from 1 to 6 years of age. Longitudinal analyses were used to evaluate the growth trajectories of offspring body mass index (BMI). Multivariate logistic regression was used to assess the effects of two adverse pregnancy outcomes, gestational diabetes mellitus (GDM) and being large-for-gestational age (LGA), on childhood overweight. Offspring of mothers with GDM and LGA has a higher annual BMI z-score from 1 to 6 years of age (all P P P P = 0.0930). Offspring of mothers with GDM and LGA had a higher BMI z-score and increased risk for overweight. Indeed, intrauterine exposure to maternal GDM and LGA could bias offspring to overweight, whereas maternal pre-pregnancy BMI may play a key role in offspring overweight for children born to mothers with GDM and LGA.

Autoimmune diabetes is a disorder of immune homeostasis that leads to targeted insulin-secreting islet β cell destruction characterized by insulitis. Human amylin (hA) is an important neuroendocrine hormone co-secreted with insulin by pancreatic β cells. Here, we report hA immune-modulatory action through inducing regulatory T cells. We ex vivo-treated human peripheral blood mononuclear cells (hPBMCs) with hA for 24 h and counted CD4+Foxp3+ regulatory T cells (Treg) using flow cytometry. Diabetic status was monitored and splenic Treg were measured in non-obese diabetic (NOD) male mice. NOD mice were intraperitoneally injected once daily with hA (n = 25) or solvent for control (n = 25) for 7 months continuously. Spleen tissues were collected at the end of intervention and processed for flow cytometry and Western blot. We found a 2.9-fold (p

Autoimmune diseases (ADs) are primarily mediated by the failure of immunological self-tolerance. Regulatory T cells (Tregs) play a critical role in the maintenance of induced tolerance to peripheral self-antigens, suppressing immoderate immune responses deleterious to the host and preventing the AD development. Tregs and suppressive cytokines are homeostatic with effective cells plus pro-inflammatory cytokines in healthy hosts which is defined as "Yang", and ADs are usually induced in case of disturbed homeostasis, which is defined as "Yin". Indeed, the Yin-Yang balance could explain the pathogenic mechanism of ADs. Tregs not only suppress CD4+ and CD8+ T cells but also can suppress other immune cells such as B cell, natural killer cell, DC and other antigen-presenting cell through cell-cell contact or secreting suppressive cytokines. In Tregs, Foxp3 as an intracellular protein displays a more specific marker than currently used other cell-surface markers (such as CD25, CD40L, CTLA-4, ICOS and GITR) in defining the naturally occurring CD4+ Tregs. Though the precise mechanism for the opposite effects of Tregs has not been fully elucidated, the importance of Tregs in ADs has been proved to be associated with kinds of immunocytes. At present, the surface marker, frequency and function of Tregs existed conflicts and hence the Tregs therapy in ADs faces challenges. Though some success has been achieved with Tregs therapy in few ADs both in murine models and humans, more effort should paid to meet the future challenges. This review summarizes the progress and discusses the phenotypic, numeric and functional abnormalities of Tregs and is the first time to systematically review the progress of Tregs therapy in kinds of ADs.

This report aimed to explore the association between the change of circulating interleukin-6 (IL-6) in patients and the development of type 1 diabetes mellitus (T1DM).Four databases (PubMed, CNKI, WanFang and Civip) were used to search and list all clinical case-control studies about serum IL-6 level in T1DM patients between Jan 1, 2000 and Aug 31, 2016.A total of 20 case-control studies with 1238 T1DM patients and 742 healthy controls were included in this study. Compared to healthy controls, the serum content of IL-6 in patients with T1DM was significantly greater (overall: SMD, 1.49; 95% CI, 1.04 to 1.93; p0.001), and notably increased in all subgroup with different age, ethnic and disease duration (all p0.001). Furthermore, the analysis in subgroup exhibited that serum levels of IL-6 in the age greater than 20-year old (SMD, 1.64; 95% CI, 0.57-2.71; p0.001), the diseased duration among 0-10years (SMD, 2.43; 95% CI, 1.42-3.44; p0.001) and the sorted American group (SMD, 1.68; 95% CI, 0.85-2.51; p0.001) were higher than those in control groups.Patients with T1DM were found to be linked to elevated level of serum IL-6, which the age, ethnic and disease durations in T1DM patients had no effect on the serum IL-6 levels for promoting diabetes mellitus.

Background: Elderly mothers are increasingly in China. We are aims to explore whether association between pregnancy to delivery interval and adverse pregnancy outcomes is affected by maternal age. Methods: A population-based retrospective study was performed in Xiamen, China. Data were derived from the Medical Birth Registry of Xiamen from 2011 to 2018. Multivariable logistic regression was used to conduct multivariable analyses based on adjusting the factors to evaluate the effect of maternal age on pregnancy outcomes. Results: Among 77,859 pregnant women with specific age, gestational diabetes mellitus (GDM) risk were increased for women aged 40 years or older (42.9%; aRR, 3.84 (3.26-4.51); P < 0.001), but not for pregnant women aged less than 25 years (9.2%; aRR, 0.60 (0.54-0.68)). Increased cesarean, preterm birth, large-for-gestational age (LGA), and low birth weight risks were more pronounced for pregnant women aged 40 years or older (66.5%, 8.4%, 26.5%, and 6.7%, respectively; aRR, 3.77 (3.14-4.52), 1.26 (0.90-1.79), 1.31 (1.08-1.60), and 1.10 (0.74-1.65), respectively; all P < 0.001). Risk of Apgar < 7 at 5 minutes were increased for women < 25 years old compared with women 35 to 39 years old (0.2% vs. 0.1%; aRR, 2.05 (0.85-4.93) vs. 0.63 (0.14-2.86)). Conclusion: Advanced maternal age increased risk of adverse pregnancy outcomes. The risk ratio of GDM, cesarean, preterm birth, LGA, and low birth weight is higher, which suggests pregnant women should guard against related risk factors and choose an appropriate mode of production. Furthermore, pregnant women should choose an ideal age for pregnancy to make themselves and child healthy.

Background: Hepatitis B surface antigen (HBsAg) positive leads to pregnancy outcomes and effects on offspring BMI z-score remains unclear in China. We are aim to explore if there is an association among HBsAg positive, BMI z-score, and pregnancy outcomes. Methods: We extracted the characteristic information and pregnancy outcomes of pregnant women, and BMI z-score from 1 to 3 years in offspring from the Medical Birth Registry of Xiamen that registered between 1 March 2011 and 30 March 2018. Results: Pregnant women with HBsAg positive had a higher risk of gestational diabetes mellitus (GDM) (1.128, 1.032-1.232), and cesarean delivery (1.116, 1.026-1.213). Interestingly, there is no difference between HBsAg positive and offspring BMI z-score (all P > 0.05). Additionally, the GDM women with HBsAg positive had higher incidence of low-for-gestation age (LGA), preterm birth, and cesarean delivery (22.95%, 9.48%, and 47.88%, respectively) (all P < 0.001). Conclusions: HBsAg positive may be a risk factor of GDM and cesarean delivery. However, HBsAg positive has no effects on offspring BMI z-score. Therefore, women should perform GDM and HBsAg screening before or during pregnancy. As well, a larger follow-up population study should be performed to confirm the association between HBsAg positive and offspring BMI z-score.

Our aim is to assess the optimal cutoff value of fasting plasma glucose (FPG) in Chinese women at 24–28 weeks’ gestation by performing oral glucose tolerance test (OGTT) to improve diagnostic rate of gestational diabetes mellitus (GDM). Data were derived from the Medical Birth Registry of Xiamen. A FPG cutoff value of 5.1 mmol/L confirmed the diagnosis of GDM in 4,794 (6.10%) pregnant women. However, a FPG cutoff value of 4.5 mmol/L should rule out the diagnosis of GDM in 35,932 (45.73%) pregnant women. If we use this cutoff value, the diagnosis of GDM to about 27.3% of pregnant women will be missed. Additionally, a 75-g OGTT was performed in pregnant women with FPG values between 4.5 and 5.1 mmol/L, avoiding the performance of formal 75-g OGTT in about 50.37% pregnant women. Meanwhile, according to maternal age and pre-pregnancy BMI categories, with FPG values between 4.5 mmol/L and 5.1 mmol/L, which had high sensitivity, to improve the diagnostic rate of GDM in all groups. Further researches are needed to present stronger evidences for the screening value of FPG in establishing the diagnosis of GDM in pregnant women.

Association between ACE gene I/D polymorphism and the risk of overweight/obesity remains controversial. We investigated the possible relationship between ACE gene I/D polymorphism and obesity in Chinese type 2 diabetes mellitus (T2DM) patients. In this study, obesity was defined as a body mass index (BMI) value ≥ 25 kg/m2and subjects were classified into 4 groups (lean, normal, overweight, and obese). PCR (polymerase chain reaction) was used to detect the ACE gene I/D polymorphism in T2DM patients. Metabolic measurements including blood glucose, lipid profile, and blood pressure were obtained. Frequencies of the ACE genotypes (DD, ID, and II) were not significant among the 4 groups of BMI-defined patients (P=0.679) while ACE II carriers showed higher systolic blood pressure (SBP) and pulse pressure (PP) (allP<0.050). Hyperglycemia, hypertension, and dyslipidemia in these T2DM patients were found to be significantly associated with BMI. In conclusion, the relationship of ACE gene I/D polymorphism with obesity is insignificant in Chinese patients with T2DM. SBP and PP might be higher in the ACE II carriers than in the DD and ID carriers.

Aims: This study aims to investigate the association between renin-angiotensin system gene polymorphism and diabetic retinopathy (DR) in Chinese patients with type 2 diabetes. Methods: We consecutively included 1491 patients for the assessment of ACE I/D and AGT M/T gene polymorphisms in 345 DR cases and 1146 patients without retinopathy (DNR). Albuminuria was defined by urine albumin creatinine ratio and albumin excretion rate. Results: Compared with the NDR patients, the DR cases displayed a higher proportion of diabetic nephropathy (32.68% vs. 6.52%, χ2 = 150.713, p < 0.001). The DR cases and DNR individuals did not differ in the frequency of genotypes and alleles of ACE I/D and AGT M/T (all p > 0.05). Intriguingly, DR patients with obesity showed higher frequency of DD (χ2 = 4.181, p = 0.041), but no significant difference exists in the other stratified BMI and hypertension analyses (all p > 0.05). Binary logistic regression displays that the association of the ACE and AGT gene polymorphisms in DR patients is not significant after adjusting for confounding covariates in all the comparisons. Conclusions: The ACE and AGT gene polymorphisms are not associated with the progress of diabetes developing into retinopathy in Chinese patients with type 2 diabetes. However, more investigations are needed to further prove the association.

We are aimed to systematically assess the worldwide trend in incidence of childhood type 1 diabetes mellitus (CT1DM) from 1965 to 2012 and to discuss whether climate affect incidence of CT1DM. We searched the relevant literatures in detail to judge the effect of different climates on incidence of CT1DM. The climates included Mediterranean, monsoon, oceanic, continental, savanna, and rainforest. According to different climates, we further researched relevant factor such as sunshine durations and latitudes. The overall incidence of CT1DM in 72 countries was 11.43 (95% CI 10.31–12.55) per 100,000 children/yr. The incidence of CT1DM in Oceanic climate [10.56 (8.69–12.42)] is highest compared with other climates; the incidence in 40°–66°34′N/S [14.71 (12.30–17.29)] is higher than other latitude groups; the incidence in sunshine durations with 3–4 hours per day [15.17 (11.14–19.20)] is highest compared with other two groups; the incidence of CT1DM from 2000 to 2012 [19.58 (14.55–24.60)] is higher than other periods; all p

// Yong-Chao Qiao 1,2 , Wei Ling 1 , Yan-Hong Pan 1 , Yin-Ling Chen 1 , Dan Zhou 1 , Yan-Mei Huang 1 , Xiao-Xi Zhang 1,3 and Hai-Lu Zhao 1,2,3 1 Center of Diabetic Systems Medicine, Guangxi Key Laboratory of Excellence, Guilin Medical University, Guilin, China 2 Department of Immunology, Xiangya School of Medicine, Central South University, Changsha, China 3 Department of Immunology, Faculty of Basic Medicine, Guilin Medical University, Guilin, China Correspondence to: Hai-Lu Zhao, email: // Keywords : tType 1 diabetes mellitus; pramlintide; postprandial glucose; adverse events; meta-analysis Received : January 20, 2017 Accepted : March 02, 2017 Published : March 08, 2017 Abstract Aims: We aim to assess the efficacy and safety of pramlintide plus insulin therapy in patients with type 1 diabetes. Methods: We included clinical studies comparing pramlintide plus insulin to placebo plus insulin. Efficacy was reflected by glycemic control and reduction in body weight and insulin use. Safety concerns were hypoglycemia and other adverse events. Subgroup analysis was performed for different doses (30, 60, 90 µg/meal) and durations (≤4, 26, 29, >29 weeks) of the treatment. Results: A total of 10 randomized placebo-controlled studies were included for this meta-analysis (pramlintide, n=1978; placebo, n=1319). Compared with controls, patients given pramlintide had significantly lower HbA1c ( p < 0.001), total daily insulin dose ( p = 0.024), mean mealtime insulin dose ( p < 0.001), body weight ( p < 0.001) and postprandial glucose level ( p = 0.002). The addition of pramlintide increased the incidence of nausea ( p < 0.001), vomiting ( p < 0.001), anorexia ( p < 0.001) and hypoglycemia ( p < 0.05) at the initiation of the treatment. The efficacy and adverse reactions of pramlintide were largely significant for the different doses and durations of the treatment. Conclusions: The addition of pramlintide to insulin therapy in patients with type 1 diabetes improves glycemic control and reduces insulin requirement and body weight while bringing transient hypoglycemia and digestive disorders.

Objectives The aim of this study was to investigate whether the concentrations of serum tumor necrosis factor-α (TNF-α), a pro-inflammatory cytokine, increased in type 2 diabetes mellitus (T2DM) and type 2 diabetic nephropathy (T2DN) patients. Methods The four databases (PubMed, CNKI, WanFang and Chinese-Cqvip) were searched from Jan 1, 1999 to October 1, 2016 for all clinical case-control studies about the serum TNF-α concentrations in T2DM and T2DN patients. All relevant data were extracted from published reports. The meta-analysis was performed to compare the changes of serum TNF-α concentrations of T2DN and T2DM patients in Eastern and Western with healthy controls. We further evaluated concentrations of serum TNF-α in T2DN patients with mincroalbuminuria or macroalbuminuria. Random-effects models were adopted to assess the pooling data among various variations. Results In total of 6 studies (744 patients and 277 healthy controls) were included in this study. Compared with healthy controls (both p 0.01), the groups of different albuminuria levels and ethnicities both showed that the serum TNF-α levels were significantly elevated in T2DN patients as well as in eastern T2DN patients ( p = 0.001), but not significant changed in western T2DN patients ( p = 0.081). The results were stable through sensitivity analysis and no significant publications bias existed in this meta-analysis. Conclusions Serum TNF-α concentrations are obviously increased in T2DN and T2DM patients, but higher in T2DN patients, suggesting an elevated inflammatory burden in T2DN patients.

To investigate the role of pannexin 1 channels in cisplatin-induced apoptosis in I-10 cells and the mechanisms.MTT assay was used to assess the cytotoxicity of cisplatin (DDP) in I-10 cells. Annexin V/PI double staining and Hoechst 33258 fluorescence staining were employed to detect early- and late-stage apoptosis of the cells, respectively. Extracellular ATP level and intracellular IP3 level in the cells were detected using commercial detection kits.I-10 cells exposed to both CBX (a pannexin 1 channel inhibitor) and DDP showed a higher cell viability compared with the cells exposed to DDP alone (P0.01). CBX significantly decreased cisplatin-induced early-stage apoptosis (P0.001) and late-stage apoptosis (P0.01), and cause obvious reductions in extracellular ATP and intracellular IP3 levels during cisplatin-induced apoptosis (P0.05).Pannexin 1 channels participate in cisplatin-induced apoptosis in I-10 cells possibly through the ATP/IP3 pathway.

Telomerase is a specialized reverse transcriptase responsible for synthesizing telomeric DNA at the ends of chromosomes. Six subunits composing the telomerase complex have been cloned: hTR (human telomerase RNA), TEP1 (telomerase-associated protein 1), hTERT (human telomerase reverse transcriptase), hsp90 (heat shock protein 90), p23, and dyskerin. In this study, we investigated the role of each the telomerase subunit on the activity of telomerase. Through down- or upregulation of telomerase, we found that only hTERT expression changed proportionally with the level of telomerase activity. The other components, TEP1, hTR, hsp90, p23, and dyskerin remained at high and unchanged levels throughout modulation. In vivo and in vitro experiments with antisense oligonucleotides against each telomerase component were also performed. Telomerase activity was decreased or abolished by antisense treatment. To correlate clinical sample status, four pairs of normal and malignant tissues from patients with oral cancer were examined. Except for the hTERT subunit, which showed differential expression in normal and cancer tissues, all other components were expressed in both normal and malignant tissues. We conclude that hTERT is a regulatable subunit, whereas the other components are expressed more constantly in cells. Although hTERT has a rate-limiting effect on enzyme activity, the other telomerase subunits (hTR, TEP1, hsp90, p23, dyskerin) participated in full enzyme activity. We hypothesize that once hTERT is expressed, all other telomerase subunits can be assembled to form a highly active holoenzyme.

Rationale Latent autoimmune diabetes in adults (LADA) refers to an autoimmune disorder characterized with detectable islets antibodies in the early diagnosis and increased autoimmune beta-cell failure progression. Notably, this kind of diabetes seems to be confused with other phenotypic diabetes. Patient concerns A young woman suffered an emotional exhaustion-induced LADA, showing asthenia, polydipsia, polyuria, and visible weight loss. The patient emotionally ended a 14-year romantic relationship, leading to the emotional flooding. Diagnoses The data from physical examination and laboratory tests exhibited as follows: glutamic acid decarboxylase antibody (GADA) = 63.83 U/mL, the fasting blood glucose (FBG) = 13.3 mmol/L, and glycated haemoglobin (HbA1c) = 10.9%. According to levels of GADA, the patient was diagnosed as LADA. Interventions The patient was clinically treated with insulin for 3-month. Then, running, diet-control, and emotional treatment were combined, such as the patient started a new relationship. Outcomes An emotional recovery initiated from a new romantic relationship and a baby, showing normal levels of GAD65 (27.007 IU/mL) and FBG (5.46) mmol/L. Lessons The emotional exhaustion might play a significant role in induction of LADA. It is important that individuals should maintain optimism, cheer, and a positive attitude.

Objective The aim of this study was used meta-analysis to investigate changes of serum tumor necrosis factor-alpha (TNF-α) in patients with type 1 diabetes mellitus (T1DM). Methods Relevant literatures were identified from PubMed, Cochrane Library, CNKI, WanFang and Chinese-Cqvip databases (published from January 1, 1999 to September 30, 2016). Eligible reports were included for pooled analysis of serum TNF-α level and subgroup analysis was performed in relation with age, disease duration and ethnicity. Results A total of 23 articles (1631 T1DM cases, 1429 healthy controls) were included for this meta-analysis. Compared with the controls, the patients had significantly increased serum TNF-α level (P < 0.001). Similar results were also found among all subgroup analysis of different age, disease duration and ethnicity (with the exception of Asian) (all P < 0.05). Regression analysis indicated that age (P = 0.680), disease duration (P = 0.957), and ethnicity (P = 0.526) of patients were not significant impact factors for the high heterogeneity. The results were stable according to the sensitivity analysis and no publication bias existed in this meta-analysis. Conclusions Serum TNF-α level in T1DM patients has significantly elevated among all age, disease duration and ethnicity groups.

Background: The existing evidence indicates increased levels of transforming growth factor beta 1 (TGF-β1) in patients with type 2 diabetes mellitus (T2DM) and those with type 2 diabetic nephropathy (T2DN); yet no meta-analysis displays a reliable result. Here we conducted a meta-analysis to evaluate characteristic changes of TGF-β1 in T2DM and diabetic nephropathy. Methods: A systematic search was conducted for eligible studies, which reported the association of TGF-β1 withT2DM and T2DN patients, in PubMed, Wangfang, Chinese-Cqvip, and China National Knowledge Infrastructure database, from February 1, 1991 to December 15, 2015. The association of serum and urine TGF-β1 in T2DM and T2DN patients should be evaluated in case-control studies. The Newcastle-Ottawa Scale was used to access the quality of the included studies, and pooling data were synthesized as standard mean difference (SMD) and 95% confidence interval (CI). The collected data were synthesized according to Cochrane Handbook for Systematic Reviews criteria. Subgroup analysis was conducted by albuminuria and ethnicity. Regression analysis and sensitivity analysis were used to explore the sources of heterogeneity. Publication bias was judged by the Egger test. Results: Sixty-three case-control studies of 364 T2DM patients (1604 T2DN patients) and 2100 healthy controls were included for meta-analysis. Compared with the controls, the cases had increased TGF-β1 levels in both serum (T2DM: SMD 1.78 μg/L; 95% CI 0.98–2.59, P

Treatment of small cells derived from the basal layer of bovine esophageal epithelium, with Triton X-100, urea and sonication resulted in a nuclear residue that was used as an immunogen for generation of monoclonal antibodies directed against nuclear components. One such antibody, designated W1, was found to label the nuclei of all cells examined. In interphase cells, the target antigen of antibody W1 was diffusely distributed in the nucleus. During metaphase, however, the W1 antigen formed prominent crescents at the poles of the mitotic spindle, diminished gradually in anaphase, and finally redistributed into the regenerating daughter nuclei. Western blotting with antibody W1 yielded a prominent polypeptide of M(r) approximately 230,000. The amino acid sequence, deduced from the nucleotide sequence of several overlapping cDNA clones that span the entire coding region, revealed that the W1 polypeptide was identical to the Nuclear Mitotic Apparatus (NuMA) protein, with a long alpha-helical central core flanked by two nonhelical domains. Interestingly, most cDNA sequences were identical to each other, except for six sequence blocks which were either inserted or deleted in individual cDNA clones. Analysis of the cDNA sequences of various clones, coupled with polymerase chain reaction amplification of cellular mRNA and genomic Southern blotting with region-specific probes, all indicated that multiple mRNA species were present in U-251 human glioma cells, derived from alternative splicing of the RNA transcript from a single NuMA/W1 gene. Besides the predominant form of the mRNA giving rise to the polypeptide of M(r) approximately 230,000, two other forms of mRNA, which arise as a result of alternative splicing and which use different translation termination codons, may yield lower molecular weight polypeptide products. Consistent with this notion, polypeptides of M(r) approximately 195,000 and approximately 194,000 have been observed in this and other studies on the NuMA/W1 protein. These data suggest that multiple isoforms of the NuMA polypeptides generated by alternative mRNA splicing may play some important functions which remain to be characterized.

Telomerase is a specialized reverse transcriptase responsible for synthesizing telomeric DNA at the ends of chromosomes. Six subunits composing the telomerase complex have been cloned: hTR (human telomerase RNA), TEP1 (telomerase-associated protein 1), hTERT (human telomerase reverse transcriptase), hsp90 (heat shock protein 90), p23, and dyskerin. In this study, we investigated the role of each the telomerase subunit on the activity of telomerase. Through down- or upregulation of telomerase, we found that only hTERT expression changed proportionally with the level of telomerase activity. The other components, TEP1, hTR, hsp90, p23, and dyskerin remained at high and unchanged levels throughout modulation. In vivo and in vitro experiments with antisense oligonucleotides against each telomerase component were also performed. Telomerase activity was decreased or abolished by antisense treatment. To correlate clinical sample status, four pairs of normal and malignant tissues from patients with oral cancer were examined. Except for the hTERT subunit, which showed differential expression in normal and cancer tissues, all other components were expressed in both normal and malignant tissues. We conclude that hTERT is a regulatable subunit, whereas the other components are expressed more constantly in cells. Although hTERT has a rate-limiting effect on enzyme activity, the other telomerase subunits (hTR, TEP1, hsp90, p23, dyskerin) participated in full enzyme activity. We hypothesize that once hTERT is expressed, all other telomerase subunits can be assembled to form a highly active holoenzyme.