GATA3 interacts with and stabilizes HIF-1α to enhance cancer cell invasiveness.

This corrects the article DOI: 10.1038/onc.2017.8.

GATA3 interacts with and stabilizes HIF-1α to enhance cancer cell invasiveness.

GATA binding protein 3 (GATA3) is indispensable in development of human organs. However, the role of GATA3 in cancers remains elusive. Hypoxia inducible factor (HIF)-1 plays an important role in pathogenesis of human cancers. Regulation of HIF-1α degradation is orchestrated through collaboration of its interacting proteins. In this study, we discover that GATA3 is upregulated in head and neck squamous cell carcinoma (HNSCC) and is an independent predictor for poor disease-free survival. GATA3 promotes invasive behaviours of HNSCC and melanoma cells in vitro and in immunodeficient mice. Mechanistically, GATA3 physically associates with HIF-1α under hypoxia to inhibit ubiquitination and proteasomal degradation of HIF-1α, which is independent of HIF-1α prolyl hydroxylation. Chromatin immunoprecipitation assays show that the GATA3/HIF-1α complex binds to and regulates HIF-1 target genes, which is also supported by the microarray analysis. Notably, the GATA3-mediated invasiveness can be significantly reversed by HIF-1α knockdown, suggesting a critical role of HIF-1α in the underlying mechanism of GATA3-mediated effects. Our findings suggest that GATA3 stabilizes HIF-1α to enhance cancer invasiveness under hypoxia and support the GATA3/HIF-1α axis as a potential therapeutic target for cancer treatment.

"I couldn't even talk to the patient": Barriers to communicating with cancer patients as perceived by nursing students.

Communication is closely related to safe practice and patient outcomes. Given that most clinicians fall into routines when communicating with patients, it is important to address communication issues early. This study explores Taiwanese nursing students' experiences of communication with patients with cancer and their families. Senior nursing students who had cared for cancer patients were recruited to participate in focus group interviews. These semi-structured interviews were recorded and transcribed for content analysis. Among the 45 participants, about 36% of them never received any communication training. Up to 76% of the participants stated that their communication with cancer patients was difficult and caused them emotional stress. Subsequent data analysis revealed four themes: disengagement, reluctance, regression and transition. Students' negative communication experiences were related to the patients' terminally ill situation; the students' lack of training, low self-efficacy and power status, poor emotional regulation, and cultural considerations. The findings of this study provide a deeper understanding of nursing students' communication experiences in oncology settings within the cultural context. Early and appropriate communication training is necessary to help students regulate their emotions and establish effective communication skills. Further studies are needed to examine the relationship among students' emotional labour, communication skills and outcomes.

Acidic FGF promotes neurite outgrowth of cortical neurons and improves neuroprotective effect in a cerebral ischemic rat model.

Acidic fibroblast growth factor (aFGF) is a neurotrophic factor which is a powerful neuroprotective and neuroregenerative factor of the nervous system. Prior study had shown that levels of FGFs significantly increase following ischemic injury, reflecting a physiological protection mechanism. However, few reports demonstrated the efficacy of applying aFGF in cerebral ischemia. A recent report showed that the intranasal aFGF treatment improved neurological functional recovery; however, it did not significantly reduce the lesion size in ischemic rats. The present study examines the neuroprotective effect of aFGF on cortical neuron-glial cultures under oxygen glucose deprivation (OGD)-induced cell damage and investigates whether epidural application of slow-released aFGF could improve benefit on ischemic stroke injury in conscious rats. We used a topical application of aFGF mixed in fibrin glue, a slow-release carrier, over the peri-ischemic cortex and examined such treatment on cerebral infarction and behavioral impairments of rats subjected to focal cerebral ischemia (FCI). Results demonstrate that aFGF effectively protected cortical neuron-glial cultures from OGD-induced neuronal damage. Neurite extension from cortical neurons was significantly enhanced by aFGF, mediated through activation of AKT and ERK. In addition, topical application of fibrin glue-mixed aFGF dose-dependently reduced ischemia-induced brain infarction and improved functional restoration in ischemic stroke rats. Slow-released aFGF not only protected hippocampal and cortical cell loss but reduced microglial infiltration in FCI rats. Our results suggest that aFGF mixed in fibrin glue could prolong the protective/regenerative efficacy of aFGF to the damaged brain tissue and thus improve the functional restorative effect of aFGF.

A functional study of proximal goat ß-casein promoter and intron 1 in immortalized goat mammary epithelial cells.

Goat ß-casein (CSN2) promoter has been extensively used to derive expression of recombinant therapeutic protein in transgenic goats; however, little direct evidence exists for signaling molecules and the cis-elements of goat CSN2 promoter in response to lactogenic hormone stimulation in goat mammary epithelial cells. Here, we use an immortalized caprine mammary epithelial cell line (CMC) to search for evidence of the above. Serial 5'-flanking regions deleted of promoter and intron 1 in goat CSN2 (-4,047 to +2,054) driven by firefly luciferase reporter gene were constructed and applied to measure promoter activity in CMC. The intron 1 region (+393 to +501) significantly decreased basal activity of the promoter. This finding contradicts other studies of the role of intron 1. The signal transducer and activator of transcription (STAT)5a played a significant role in activating promoter activity by prolactin stimulation. Hydrocortisone enhanced and prolonged the activity of STAT5a and promoter in CMC, but was independent of the glucocorticoid receptor response element. The minimum length of the CSN2 promoter segment in response to lactogenic stimulation was confirmed by 5' serial deletions. A cis-element located from -300 to -90 in proximal goat CSN2 promoter that is absent in bovine and human CSN2 promoter was newly identified. We demonstrated the presence of a STAT5a binding site (-102 to -82) and preservation of the guanosine nucleotide at position -90 based on responses to the presence of lactogenic hormone using internal deletions and point mutations of the predicted STAT5a binding site, and chromatin immunoprecipitation assay. Together, these findings demonstrate that the proximal -300 bp of goat CSN2 promoter containing the STAT5a binding site (-102 to -82) is the response element for lactogenic hormone stimulation. Additionally, intron 1 may be required for tissue or developmental stage-specific expression in mammary gland. The role of the far-distal regions of goat CSN2 promoter in high-level lactogenic hormone induction and specific expression require further examination.

ß-1,4-Galactosyltransferase III suppresses extravillous trophoblast invasion through modifying ß1-integrin glycosylation.

INTRODUCTION: Glycosylation controls diverse protein functions and regulates various cellular phenotypes. Trophoblast invasion is essential for normal placental development. However, the role of glycosylation in human placenta throughout pregnancy is still unclear. The ß-1,4-galactosyltransferase III (B4GALT3) has been found to regulate cancer cell invasion. We therefore investigated the expression of B4GALT3 in placenta and its roles in trophoblast. METHODS: B4GALT3 protein expression was examined by quantitative Western blotting analysis in human placentas. For identification of B4GALT3-positive cells in normal human placenta, immunohistochemistry and immunofluorescence methods were used. To investigate effects of B4GALT3 on extravillous trophoblast (EVT)-like cell and primary EVT cells, we analyzed cell growth, adhesion, migration, and invasion in mock and B4GALT3-transfected cell. RESULTS: B4GALT3 expression significantly increased in third trimester human placenta. Immunostaining revealed that B4GALT3 expressed in placental villous cytotrophoblast, syncytiotrophoblast, and a subpopulation of EVT cells throughout pregnancy. Interestingly, we found increases in the expression level and percentage of B4GALT3-positive cells in third trimester EVT, but not in syncytiotrophoblasts and cytotrophoblasts of placental villi. Overexpression of B4GALT3 in HTR8/SVneo cells and primary trophoblast cells significantly suppressed cell migration. In addition, B4GALT3 suppressed cell invasion, and enhanced cell adhesion to laminin in HTR8/SVneo cells. Notably, we found that B4GALT3 modified glycans on ß1-integrin, suppressed focal adhesion kinase (FAK) signaling, and enhanced ß1-integrin degradation. DISCUSSION: We propose that B4GALT3-mediated glycosylation change not only enhances ß1-integrin binding to laminin, but also attenuates ß1-integrin stability. Our findings suggest that B4GALT3 is a critical regulator for suppressing EVT invasion in the late stages of pregnancy.

[Decellularized collagen matrix from tilapia fish scales for corneal reconstruction (BioCornea)]. / Dezellularisierte Kollagenmatrix aus der Schuppe des Tilapia-Fisches als Hornhautersatz ("BioCornea").

BACKGROUND: The worldwide need for donor corneal tissue clearly exceeds the availability of transplantable human tissue; therefore, recent efforts aim to identify and characterize alternative tissues, such as decellularized collagen scaffolds. OBJECTIVES: The transparent fish scales of tilapia (Oreochromis mossambicus) were analyzed as a potential alternative for corneal reconstruction ("BioCornea"). MATERIAL AND METHODS: The article gives a review of the literature and own preliminary results. After decellularization the tissue characteristics of the fish scales, the repopulation with corneal epithelium and stromal cells, immunogenicity, the feasibility of corneal transplantation and the angiogenic properties were analyzed in vitro and in various animal models. RESULTS: The fish scales mainly consist of collagen type I and show an architecture that is similar to the human cornea. Corneal epithelium and stromal cells are able to grow over and into the scaffold. It is possible to transplant fish scales in various animal models without severe inflammatory responses. Furthermore, in mice, less blood and lymphatic vessels grow into the xenograft when compared to conventional allogenic transplants. CONCLUSION: Preliminary results with decellularized tilapia fish scales as an alternative for corneal reconstruction ("BioCornea") are promising.

Identification of novel loci for bipolar I disorder in a multi-stage genome-wide association study.

OBJECTIVE: Identification of genetic variants that influence bipolar I disorder (BPD-I) through genome-wide association (GWA) studies is limited in Asian populations. The current study aimed to identify novel common variants for BPD-I in an ethnically homogeneous Taiwanese sample using a multi-stage GWA study design. METHOD: At the discovery stage, 200 BPD-I patients and 200 controls that combined to form 16 pools were genotyped with 1 million markers. Utilizing a newly developed rank-based method, top-ranked markers were selected. After validation with individual genotyping, a fine-mapping association study was conducted to identify associated loci using 240 patients and 240 controls. At the last stage, independent samples were collected (351 cases and 341 controls) for replication. RESULTS: Among the top-ranked markers from the discovery stage, eight genes and 15 individual SNPs were evaluated in the fine-mapping stage. At this stage, rs7619173, which is not in a gene coding region, showed the most significant association (P = 2 ∗ 10(-5)) with BPD-I. Four genes had empirical P-values<0.05, including KCNH7 (P = 0.0047), MYST4 (P = 0.0047), NRXN3 (P = 0.0095), and SEMA3D (P = 0.037). For markers genotyped in replication samples, rs7619173 exhibited a significant association (P(combined) = 2 ∗ 10(-4)) after multiple testing correction, while markers rs11001178 (MYST4) and rs2217887 (NRXN3) showed weak associations (P(combined) = 0.02) with BPD-I. CONCLUSION: A multi-stage GWA design has the potential to uncover the underlying pathogenesis of a complex trait. Findings in the present study highlight three loci that warrant further investigation for bipolar.

Expression of GALNT2 in human extravillous trophoblasts and its suppressive role in trophoblast invasion.

Extravillus trophoblast (EVT) invasion plays a critical role in placental development. Integrins bind to extracellular matrix (ECM) proteins to mediate EVT cell adhesion, migration, and invasion. Changes in O-glycans on ß1-integrin have been found to regulate cancer cell behavior. We hypothesize that O-glycosyltransferases can regulate EVT invasion through modulating the glycosylation and function of ß1-integrin. Here, we found that the GALNT1 and GALNT2 mRNA were highly expressed in HTR8/SVneo and first trimester EVT cells. Immunohistochemstry and immunofluorescence staining showed that GALNT2 was expressed in subpopulations of EVT cells in deciduas, but not in syncytiotrophoblasts and cytotrophoblasts of placental villi. The percentage of GALNT2-positive EVT cells increased with gestational ages. Overexpression of GALNT2 in HTR8/SVneo cells significantly enhanced cell-collagen IV adhesion, but suppressed cell migration and invasion. Notably, we found that GALNT2 increased the expression of Tn antigen (GalNAc-Ser/Thr) on ß1-integrin as revealed by Vicia Villosa agglutinin (VVA) binding. Furthermore, GALNT2 suppressed the phosphorylation of focal adhesion kinase (FAK), a crucial downstream signaling molecule of ß1-integrin. Our findings suggest that GALNT2 is a critical initiating enzyme of O-glycosylation for regulating EVT invasion.

Regulation of chondroitin sulphate proteoglycan and reactive gliosis after spinal cord transection: effects of peripheral nerve graft and fibroblast growth factor 1.

AIMS: The combined treatment of peripheral nerve (PN) graft and fibroblast growth factor (FGF)-1 for spinal cord injury produces functional recovery, but how it affects injury events is still unknown. This project studied the effect of PN graft and FGF-1 on white matter degeneration following spinal cord injury. METHODS: Rats were divided into four groups: (i) complete spinal cord transection and T8 segment removed; the remaining three groups underwent transection followed by (ii) PN grafting; (iii) supply of exogenous FGF-1; and (iv) PN grafting plus FGF-1 treatment. Chondroitin sulphate proteoglycan (CSPG) deposition, astrocytes and macrophage activation, cavity size, and calcitonin gene-related peptide and synaptophysin immunoreactivity were compared. RESULTS: Peripheral nerve grafting increased CSPG levels compared to transection surgery alone. This CSPG was associated with the proximity to the PN graft. FGF-1 reduced CSPG deposition in grafted animals regardless of the proximity to the graft. The CSPG reduction was accompanied by reduced GFAP expression and macrophage activation. The amount of CSPG with dissociated glycosaminoglycan did not differ between groups. FGF-1 in Schwann cell-astrocyte coculture did not reduce CSPG deposition. Furthermore, the PN graft increased the calcitonin gene-related peptide immunoreactivity and altered the distribution of synaptophysin-positive axons. CONCLUSION: Peripheral nerve graft supported sensory re-innervation and partial protection of the grey matter, but up-regulated CSPG in the graft-stump junction compared to non-grafted rats. The reduction of CSPG was caused by FGF-1-PN synergy, and did not involve dissociation of CSPG or the suppression of a general immune response.

Cigarette smoking and proteinuria in Taiwanese men with Type 2 diabetes mellitus.

AIMS: Cigarette smoking is a well-known risk factor associated with diabetic nephropathy. The objective of this study was to further investigate the dose-response effect of tobacco exposure on proteinuria in males with Type 2 diabetes. METHODS: Five hundred and nine males with Type 2 diabetes were selected from a cohort participating in a glucose control study in Taiwan. Pack-years of cigarette smoking were calculated to define tobacco exposure. Proteinuria was identified if albumin-to-creatinine ratio was > or = 30 mg/g in at least two of three consecutive urine tests. Logistic regression and trend tests were used to delineate the association between smoking status and proteinuria. RESULTS: Compared with non-smokers, those who had smoked 15-30 or more than 30 pack-years were respectively 2.78 (95% CI 1.34-5.76, P < 0.01) and 3.20 (95% CI 1.74-5.86, P < 0.001) times more likely to develop proteinuria. The dose-response effect of tobacco exposure on the development of proteinuria is highly significant in all subjects (P = 0.001) and in subgroups with relatively short duration of diabetes mellitus (P < 0.001), good blood pressure control (P = 0.001) and those of young age (P = 0.007). CONCLUSIONS: The current study shows a clear dose-response effect of cigarette smoking on development of proteinuria in male Type 2 diabetic patients. These findings reinforce the urgent need to encourage diabetic patients to stop smoking regardless of age, duration of diabetes mellitus or status of blood pressure control.

The influence of donor and recipient factors in allograft rejection of the human cornea.

UNLABELLED: The health of the corneal endothelium is essential in maintaining the clarity of the transplanted human cornea. Immune-mediated endothelial rejection is a complex series of events, which may culminate in the decompensation of the donor button. It is the commonest instigator of failure in penetrating corneal transplantation. METHODS: This retrospective case note review of 203 penetrating keratoplasties with adequate follow-up data during a 5-year study period from 1 January 2000 to 31 December 2003 at Manchester Royal Eye Hospital, were used for analysis. RESULTS: Forty-three of the 203 donor grafts underwent at least one single episode of immune-mediated endothelial rejection, an incidence of 21% over a 5-year follow-up. Recipient's age was inversely associated with the risk of rejection. The average age for the cohort of 58.7 years and average age for rejecting patients of 47.6 years were strongly significantly different (P=0.009). Rejection in keratoconic patients accounted for 30% of cases. Death to enucleation time (P=0.03) was also associated with an increased risk of rejection. CONCLUSION: Although penetrating keratoplasty is an effective long-term treatment option for improving visual function, the endothelial rejection rate in our study was 21% over a mean follow-up of over 5 years. Host vascularisation, regrafts, younger recipient age group, and donor factors were found to be significantly associated with a risk of rejection. Rejection in keratoconic recipients was more common than expected.

Prevalence of metabolic syndrome among patients with schizophrenia or schizoaffective disorder in Taiwan.

OBJECTIVE: Because of ethnic differences in metabolic syndrome (MS) criteria, this study aimed to investigate the MS prevalence among patients with schizophrenia or schizoaffective disorder in Taiwan. METHOD: We recruited 650 patients with schizophrenia or schizoaffective disorder from 36 psychiatric institutions. The MS prevalence was assessed based on the modified Adult Treatment Panel (ATP) III criteria for Asians. RESULTS: The overall MS prevalence was 34.9%, with 38.9% in female and 31.5% in male patients respectively. The difference of MS prevalence between our sample and the general population was marked in male patients under 40 years of age and in female patients under 50 years old. Body mass index > or =24 and age over 40 years old are two important risk factors of MS. Female and polypharmacy had marginal significance with the presence of MS. CONCLUSION: Patients with schizophrenia or schizoaffective disorder in Taiwan had a high prevalence of MS, which appeared early in their lives.

Expressed transcripts associated with high rates of egg production in chicken ovarian follicles.

The purpose of this study was to characterize differentially expressed transcripts associated with varying rates of egg production in Taiwan country chickens. Ovarian follicles were isolated from two strains of chicken which showed low (B) or high (L2) rates of egg production, then processed for RNA extraction and cDNA library construction. Three thousand and eight forty clones were randomly selected from the cDNA library and amplified by PCR, then used in microarray analysis. Differentially expressed transcripts (P<0.05, log(2)> or = 1.75) were sequenced, and aligned using GenBank. This analysis revealed 20 non-redundant sequences which corresponded to known transcripts. Eight transcripts were expressed at a higher level in ovarian tissue prepared from chicken strain B, and 12 transcripts were expressed at a higher level in L2 birds. These differential patterns of expression were confirmed by semi-quantitative RT-PCR. We show that transcripts of cyclin B2 (cycB2), ferritin heavy polypeptide 1 (FTH1), Gag-Pol polyprotein, thymosin beta4 (TB4) and elongation factor 1 alpha1 (EEF1A1) were enriched in B strain ovarian follicles. In contrast, thioredoxin (TXN), acetyl-CoA dehydrogenase long chain (ACADL), inhibitor of growth family member 4 (ING4) and annexin II (ANXA2) were expressed in at higher levels in the L2 strain. We suggest that our approach may lead to the isolation of effective molecular markers that can be used in selection programs in Taiwan country chickens.

C2GnT-M is downregulated in colorectal cancer and its re-expression causes growth inhibition of colon cancer cells.

Changes in carbohydrates on the cell surface are associated with tumor malignancy. The mucin-type core 2 beta-1,6-N-acetylglucosaminyltransferase (C2GnT-M) is highly expressed in the gastrointestinal tract and catalyses the formation of core 2, core 4, and blood group I branches on O-glycans. In the present study, we evaluated the role of C2GnT-M in colorectal cancer. C2GnT-M downexpression was observed in 73.6% of the primary tumors from colorectal cancer patients (39 of 53) analysed by cancer profiling array. Consistently, the majority of colon cancer cell lines and primary colon tumors expressed lower levels of C2GnT-M than did normal colon tissues by RT-PCR. HCT116 cells stably transfected with C2GnT-M inhibited expression of the core 1 structure, Galbeta1,3GalNAcalpha1-Ser/Thr, on the cell surface. Moreover, C2GnT-M expression suppressed cell adhesion, motility, and invasion as well as colony formation ability. The growth of C2GnT-M-transfected HCT116 and SW480 cells was dramatically suppressed, and the cell death induced by C2GnT-M was demonstrated by an increase in the annexin V-positive cells. Interestingly, C2GnT-M inhibited cell adhesion to collagen IV and fibronectin, and decreased tyrosine phosphorylation of paxillin, indicating that the changes in cancer behavior may be partly mediated by integrin-signaling pathways. Tumor growth in vivo was also significantly suppressed by C2GnT-M in the xenografts of nude mice. These results demonstrate that C2GnT-M is frequently downregulated in colorectal cancer and suppresses colon cancer cell growth.

Effects of glucose concentration on in vitro fertilization in BALB/c mice.

BALB/c mice are widely used in genetic, tumour and immunological studies. However, the mice demonstrate a lower reproduction rate, low fertility and small litters, because of their highly genetic homozygoisty. Based on in vitro fertilization (IVF), a routine technique for biomedical studies, it is worth to evaluate the effects to BALB/c mice on IVF efficiency. In order to test the genetic factor affecting the IVF efficiency of BALB/c, four reciprocal IVF tests of BALB/cByJ and FVB/NCrl mice were performed. The results showed that the average fertility of IVF sponsored by FVB/NCrl spermatozoa was 69.6%, but only 12.1% was obtained from BALB/cByJ strain. Effect of glucose contained in the culture medium to the IVF efficiency of BALB/cByJ was also evaluated. The results showed that the fertility of BALB/cByJ spermatozoa incubated with 0, 2.7, 5.5, 11.1 and 22.2 mm of glucose in the TYH medium were 6.8, 9.9, 13.9, 32.7 and 22.2%, respectively. It is showed that IVF efficiency of BALB/cByJ spermatozoa could be improved depending on the concentration of glucose in the IVF medium. According to the results, it is beleived that lower IVF of BALB/cByJ mice might be due to the genetic defect in spermatozoa and increasing glucose in the IVF medium which significantly affect the IVF efficiency of BALB/cByl via activating the spermatozoa.

Comparative analyses of multi-species sequences from targeted genomic regions.

The systematic comparison of genomic sequences from different organisms represents a central focus of contemporary genome analysis. Comparative analyses of vertebrate sequences can identify coding and conserved non-coding regions, including regulatory elements, and provide insight into the forces that have rendered modern-day genomes. As a complement to whole-genome sequencing efforts, we are sequencing and comparing targeted genomic regions in multiple, evolutionarily diverse vertebrates. Here we report the generation and analysis of over 12 megabases (Mb) of sequence from 12 species, all derived from the genomic region orthologous to a segment of about 1.8 Mb on human chromosome 7 containing ten genes, including the gene mutated in cystic fibrosis. These sequences show conservation reflecting both functional constraints and the neutral mutational events that shaped this genomic region. In particular, we identify substantial numbers of conserved non-coding segments beyond those previously identified experimentally, most of which are not detectable by pair-wise sequence comparisons alone. Analysis of transposable element insertions highlights the variation in genome dynamics among these species and confirms the placement of rodents as a sister group to the primates.

A metastatic pituitary carcinoid tumor successfully treated with gamma knife radiosurgery.

Intracranial metastasis occurs in a certain number of patients with carcinoid tumor. However, carcinoid tumor with metastasis to the pituitary gland is extremely rare. Up to the present, no effective treatment for either a metastatic intracranial carcinoid tumor or a metastatic pituitary lesion of any origin has been documented. We have treated a case of metastatic carcinoid tumor of the pituitary gland with transsphenoidal tumor resection followed by gamma knife radiosurgery. A 59-year-old man presented with headache and left oculomotor palsy. He was treated at the same hospital for bronchial atypical carcinoid tumor one and a half years ago. Magnetic resonance image of the brain showed a pituitary tumor. There were no signs of recurrent or metastatic lesion elsewhere despite thorough investigation. Transsphenoidal approach for removal of tumor was done and the pathology turned out to be a metastatic carcinoid tumor. Subsequent gamma knife radiosurgery was given for residual tumor. The oculomotor palsy improved after radiosurgery. No neurological deficit occurred. Follow-up CT scan of the brain showed complete resolution of the tumor. We concluded that gamma knife radiosurgery could be used to treat a metastatic intracranial carcinoid tumor. It can also be used to treat a metastatic lesion of the pituitary gland without causing neurological deficit.

Loss of mismatch repair activity in simian virus 40 large T antigen-immortalized BPH-1 human prostatic epithelial cell line.

Simian virus 40 large T antigen (SVLTAg) has been used to immortalize cells; however, the mechanism leading to immortalization is still unclear. We hypothesize that DNA mismatch repair (MMR) activity is important during SVLTAg-induced immortalization. To test this hypothesis, we used the SVLTAg-immortalized cell line BPH-1 derived from human benign prostate epithelial cells to analyze MMR activity and the expression of MMR genes (hMLH1, hPMS1, hPMS2, hMSH2, hMSH3, and hMSH6). The results demonstrated that BPH-1 cells were deficient in repairing G:T, A:C, and G:G mispairs in bacteriophage M13mp2. Reverse-transcription polymerase chain reaction experiments indicated MMR genes (hMSH3, hMSH6, and hPMS1) were expressed at a low level in BPH-1 cells. In contrast, all six MMR genes were expressed in human benign prostate hyperplasia tissues. Downregulation of hMSH3, hMSH6, and hPMS1 genes is not a result of the hypermethylation mechanism because demethylation with 5-aza-2'-deoxycytidine did not restore expression of these genes. Although the hMLH1 gene is expressed in BPH-1 cells, western blotting and exon analyses demonstrated that hMLH1 was mutated and/or deleted in BPH-1 cells.