Combined Microbiome and Metabolome Analysis Reveals a Novel Interplay Between Intestinal Flora and Serum Metabolites in Lung Cancer.

As the leading cause of cancer death, lung cancer seriously endangers human health and quality of life. Although many studies have reported the intestinal microbial composition of lung cancer, little is known about the interplay between intestinal microbiome and metabolites and how they affect the development of lung cancer. Herein, we combined 16S ribosomal RNA (rRNA) gene sequencing and liquid chromatography-mass spectrometry (LC-MS) technology to analyze intestinal microbiota composition and serum metabolism profile in a cohort of 30 lung cancer patients with different stages and 15 healthy individuals. Compared with healthy people, we found that the structure of intestinal microbiota in lung cancer patients had changed significantly (Adonis, p = 0.021). In order to determine how intestinal flora affects the occurrence and development of lung cancer, the Spearman rank correlation test was used to find the connection between differential microorganisms and differential metabolites. It was found that as thez disease progressed, L-valine decreased. Correspondingly, the abundance of Lachnospiraceae_UCG-006, the genus with the strongest association with L-valine, also decreased in lung cancer groups. Correlation analysis showed that the gut microbiome and serum metabolic profile had a strong synergy, and Lachnospiraceae_UCG-006 was closely related to L-valine. In summary, this study described the characteristics of intestinal flora and serum metabolic profiles of lung cancer patients with different stages. It revealed that lung cancer may be the result of the mutual regulation of L-valine and Lachnospiraceae_UCG-006 through the aminoacyl-tRNA biosynthesis pathway, and proposed that L-valine may be a potential marker for the diagnosis of lung cancer.

Erratum: Glycyrrhizic acid inhibits leukemia cell growth and migration via blocking AKT/mTOR/STAT3 signaling.

[This corrects the article on p. 5175 in vol. 8, PMID: 26191214.].

ROS Promote Ox-LDL-Induced Platelet Activation by Up-Regulating Autophagy Through the Inhibition of the PI3K/AKT/mTOR Pathway.

BACKGROUND/AIMS: Oxidized low-density lipoprotein (oxLDL) promotes unregulated platelet activation in patients with dyslipidemic disorders. Although oxLDL stimulates activating signaling, researchers have not clearly determined how these events drive accelerated thrombosis. Here, we describe the mechanism by which ROS regulate autophagy during ox-LDL-induced platelet activation by modulating the PI3K/AKT/mTOR signaling pathway. METHODS: For in vitro experiments, ox-LDL, the ROS scavenger N-acetylcysteine (NAC), the mTOR inhibitor rapamycin and the autophagy inhibitor 3-MA were used alone or in combination with other compounds to treat platelets. Then, platelet aggregation was evaluated on an aggregometer and platelet adhesion was measured under shear stress. The levels of a platelet activation marker (CD62p) were measured by flow cytometry, reactive oxygen species (ROS) levels were then quantified by measuring DCFH-DA fluorescence intensity via flow cytometry. Nitric oxide (NO) and superoxide (O2·-) levels were determined by the nitric acid deoxidize enzyme method and lucigenin-enhanced chemiluminescence (CL), respectively. Transmission electron microscopy was used to observe the autophagosome formation, immunofluorescence staining was employed to detect LC3 expression and western blotting was used to measure the levels of PI3K/AKT/mTOR pathway- and autophagy-related proteins. RESULTS: Ox-LDL-induced platelets showed a significant increase in platelet aggregation and adhesion, CD62p expression, ROS level and O2·- content, with an elevated LC3II/LC3I ratio and Beclin1 expression, but a dramatic reduction in the levels of p62 and pathway-related proteins (all P < 0.05). However, platelet activation and autophagy were aggravated by the Rapamycin treatment, and decreased following treatment with NAC, 3-MA, or NAC and 3-MA, together with increased activity of the PI3K/AKT/mTOR pathway. Additionally, decreased platelet activation and autophagy were observed in platelets treated with NAC and Rapamycin or Rapamycin and 3-MA compared with platelets treated with Rapamycin alone, suggesting that both NAC and 3-MA reversed the effects of Rapamycin. CONCLUSION: Inhibition of ROS production may reduce autophagy to suppress ox-LDL-induced platelet activation by activating PI3K/AKT/mTOR pathway.

The inhibition of UBC13 expression and blockage of the DNMT1-CHFR-Aurora A pathway contribute to paclitaxel resistance in ovarian cancer.

Paclitaxel is widely used as a first-line chemotherapeutic drug for patients with ovarian cancer and other solid cancers, but drug resistance occurs frequently, resulting in ovarian cancer still presenting as the highest lethality among all gynecological tumors. Here, using DIGE quantitative proteomics, we identified UBC13 as down-regulated in paclitaxel-resistant ovarian cancer cells, and it was further revealed by immunohistochemical staining that UBC13 low-expression was associated with poorer prognosis and shorter survival of the patients. Through gene function experiments, we found that paclitaxel exposure induced UBC13 down-regulation, and the enforced change in UBC13 expression altered the sensitivity to paclitaxel. Meanwhile, the reduction of UBC13 increased DNMT1 levels by attenuating its ubiquitination, and the up-regulated DNMT1 enhanced the CHFR promoter DNA methylation levels, leading to a reduction of CHFR expression, and an increased in the levels of Aurora A. Our findings revealed a novel function for UBC13 in regulating paclitaxel sensitivity through a DNMT1-CHFR-Aurora A pathway in ovarian cancer cells. UBC13 could potentially be employed as a therapeutic molecular drug for reversing paclitaxel resistance in ovarian cancer patients.

The lncRNA PDIA3P Interacts with miR-185-5p to Modulate Oral Squamous Cell Carcinoma Progression by Targeting Cyclin D2.

Long noncoding RNAs (lncRNAs) are emerging as important regulators during tumorigenesis by serving as competing endogenous RNAs (ceRNAs). In this study, the qRT-PCR results indicated that the lncRNA protein disulfide isomerase family A member 3 pseudogene 1 (PDIA3P) was overexpressed in oral squamous cell carcinoma (OSCC) and decreased the survival rate of OSCC patients. CCK-8 and clonal colony formation assays were used to detect the effects of PDIA3P on proliferation. Results revealed that silencing PDIA3P by small interfering RNA (siRNA) inhibited OSCC cell proliferation and repressed tumor growth and reduced the expression of proliferation antigen Ki-67 in vivo. Furthermore, the interaction between PDIA3P and miRNAs was then analyzed by qRT-PCR and luciferase reporter gene assay. We found that PDIA3P negatively regulated miR-185-5p in OSCC cells. Simultaneously, we found that silencing PDIA3P by siRNA suppressed proliferation via miR-185-5p in OSCC cells. Moreover, silencing PDIA3P by siRNA inhibited CCND2 protein (no influence on mRNA levels) expression via miR-185-5p in OSCC cells, and CCND2 facilitated cell proliferation of SCC4 and SCC15 cells induced by sh-PDIA3P#1. Therefore, our study demonstrated that PDIA3P may be a therapeutic target for the treatment of OSCC.

Endoplasmic reticulum stress induces autophagy and apoptosis while inhibiting proliferation and drug resistance in multiple myeloma through the PI3K/Akt/mTOR signaling pathway.

We investigated the effects of endoplasmic reticulum stress (ERS) on autophagy, proliferation, apoptosis, and drug resistance in multiple myeloma (MM). MM patients enrolled in our study (n = 268) were classified into sensitive and resistant groups based on chemotherapy efficacy, and their serum levels of ß2-MG, albumin (ALB), lactic dehydrogenase (LDH), Ca2+ and hemoglobin were determined. In addition, human MM U266 and MOLP-2/R cells were divided into blank, tunicamycin (TM), TM + insulin-like growth factor-1 (IGF-1), and TM + rapamycin groups, and measured expression of ERS-related, PI3K/Akt/mTOR pathway-related, and autophagy-related mRNA and proteins. Serum levels of ß2-MG, LDH and Ca2+, and expression of PI3K, Akt, and mTOR were higher in the resistant than sensitive group. Serum levels of ALB and hemoglobin, and expression of glucose-regulated protein 78 (GRP78), GRP94, microtubule associated protein 1 light chain 3 (LC3), and Beclin1, were lower in the resistant than sensitive group. In U266 cells treated with TM and IGF-1 or rapamycin, ERS promoted autophagy and apoptosis while inhibiting proliferation through inhibition of PI3K/Akt/mTOR signaling. ERS also reversed drug resistance in MOLP-2/R cells via the PI3K/Akt/mTOR signaling pathway. These data suggest that ERS activation could be exploited for therapeutic benefits in the treatment of MM.

Celecoxib-erlotinib combination delays growth and inhibits angiogenesis in EGFR-mutated lung cancer.

Combination treatment for non-small cell lung cancer (NSCLC) is becoming more popular due to the anticipation that it may be more effective than single drug treatment. In addition, there are efforts to genetically screen patients for specific mutations in light of attempting to administer specific anticancer agents that are most effective. In this study, we evaluate the anticancer and anti-angiogenic effects of low dose celecoxib-erlotinib combination in NSCLC in vitro and in vivo. In NSCLC cells harboring epidermal growth factor receptor (EGFR) mutations, combination celecoxib-erlotinib treatment led to synergistic cell death, but there was minimal efficacy in NSCLC cells with wild-type EGFR. In xenograft models, combination treatment also demonstrated greater inhibition of tumor growth compared to individual treatment. The anti-tumor effect observed was secondary to the targeting of angiogenesis, evidenced by decreased vascular endothelial growth factor A (VEGFA) levels and decreased levels of CD31 and microvessel density. Combination treatment targets angiogenesis through the modulation of of the PI3K/AKT and ERK/Raf1-1 pathway in NSCLC with EGFR exon 19 deletions. These findings may have significant clinical implications in patients with tumors harboring EGFR exon 19 deletions as they may be particularly sensitive to this regimen.

Glycyrrhizic acid inhibits leukemia cell growth and migration via blocking AKT/mTOR/STAT3 signaling.

Glycyrrhizic acid (GA) is the bioactive compound of licorice and has been used as an herbal medicine because of its anti-viral, anti-cancer, and anti-inflammatory properties. This study was designed to investigate the effects of GA on leukemia cells growth, migration, and the mechanisms underlying the anti-cancer activities of GA. MTT test was used to detect the effect of GA on TF-1 leukemia cell growth. Wound closure assay and Transwell were adopted to assess the effect of GA on TF-1 migration and invasion. Migration and invasion related proteins including AKT and mTOR were detected by western blot assay. We further used western blot and immunofluorescence assay to evaluate the effect of GA on STAT3 phosphorylation in vitro. We also evaluated the anti-tumor effect of GA in TF-1 tumor bearing BALB/c mice model. The present study showed GA significant inhibit of TF-1 proliferation in a dose and time-dependent manner. GA could remarkably inhibit TF-1 cell migration and invasion; meanwhile effectively suppress AKT, mTOR, and STAT3 phosphorylation in TF-1 cells. GA in 100 mg/kg/ could inhibit the tumor growth in vivo and down-regulated AKT, mTOR, and STAT3 phosphorylation in TF-1 tumor tissues. Our findings suggest that GA is a promising therapeutic agent for leukemia that targets the AKT/mTOR/STAT3 pathway.

TXNDC17 promotes paclitaxel resistance via inducing autophagy in ovarian cancer.

Paclitaxel is recommended as a first-line chemotherapeutic agent against ovarian cancer, but drug resistance becomes a major limitation of its success clinically. The key molecule or mechanism associated with paclitaxel resistance in ovarian cancer still remains unclear. Here, we showed that TXNDC17 screened from 356 differentially expressed proteins by LC-MS/MS label-free quantitative proteomics was more highly expressed in paclitaxel-resistant ovarian cancer cells and tissues, and the high expression of TXNDC17 was associated with poorer prognostic factors and exhibited shortened survival in 157 ovarian cancer patients. Moreover, paclitaxel exposure induced upregulation of TXNDC17 and BECN1 expression, increase of autophagosome formation, and autophagic flux that conferred cytoprotection for ovarian cancer cells from paclitaxel. TXNDC17 inhibition by siRNA or enforced overexpression by a pcDNA3.1(+)-TXNDC17 plasmid correspondingly decreased or increased the autophagy response and paclitaxel resistance. Additionally, the downregulation of BECN1 by siRNA attenuated the activation of autophagy and cytoprotection from paclitaxel induced by TXNDC17 overexpression in ovarian cancer cells. Thus, our findings suggest that TXNDC17, through participation of BECN1, induces autophagy and consequently results in paclitaxel resistance in ovarian cancer. TXNDC17 may be a potential predictor or target in ovarian cancer therapeutics.

Silencing of CCR7 inhibits the growth, invasion and migration of prostate cancer cells induced by VEGFC.

Early in prostate cancer development, tumor cells express vascular endothelial growth factor C (VEGF-C), a secreted molecule that is important in angiogenesis progression. CC-chemokine receptor 7 (CCR7), another protein involved in angiogenesis, is strongly expressed in most human cancers, where it activated promotes tumor growth as well as favoring tumor cell invasion and migration. The present study aimed to investigate the effect of down-regulating CCR7 expression on the growth of human prostate cancer cells stimulated by VEGFC. The CCR7-specific small interfering RNA (siRNA) plasmid vector was constructed and then transfected into prostate cancer cells. The expression of CCR7 mRNA and protein was detected by quantitative polymerase chain reaction and western blot analysis, respectively. Cell proliferation, apoptosis, cell cycle distribution and cell migration were assessed following knockdown of CCR7 by RNA interference (RNAi). Western blot analysis was used to identify differentially expressed angiogenesis- and cell cycle-associated proteins in cells with silenced CCR7. The expression levels of CCR7 in prostate cancer cells transfected with siRNA were decreased, leading to a significant inhibition of prostate cancer cell proliferation, migration and invasion induced by VEGFC. Western blot analysis revealed that silencing of CCR7 may inhibit vascular endothelial growth factor, matrix metalloproteinase (MMP)-2 and MMP-9 protein expression. In conclusion, the present study demonstrated that RNAi can effectively silence CCR7 gene expression and inhibit the growth of prostate cancer cells, which indicates that there is a potential of targeting CCR7 as a novel gene therapy approach for the treatment of prostate cancer.

[Risk factors of predicting residual disease in women with stage I a1 squamous cervical carcinoma after conization].

OBJECTIVE: To evaluate the prevalence and influencing factors of residual disease in women with stage I a1 squamous cervical carcinoma after conization. METHODS: The medical records and histopathologic slides of 83 women diagnosed with stage I a1 squamous cervical carcinoma after cervical conization undergoing subsequent hysterectomy at our hospital between January 2003 and December 2007 were reviewed. The correlations between the presence of residual lesions and clinicopathological features were analyzed. RESULTS: Among them, 31 (37.3%) had residual disease in hysterectomy specimens, including CIN1 (n = 5), CIN2-3 (n = 10), microinvasive carcinoma (n = 11) and invasive carcinoma (n = 5). In univariate analysis, menopause, procedure of conization, and status of cone margins were associated with the prevalence of residual disease in stage I a1 cervical carcinoma after conization. However, Logistic regression analysis revealed status of cone margins as an independent risk factor for residual disease in stage I a1 cervical carcinoma after conization. CONCLUSION: Status of cone margins is an independent risk factor for residual disease in stage I a1 cervical carcinoma after conization. Further treatment should be performed in patients with positive or nearing cone margins.

[Reversing paclitaxel-resistance of SKOV3-TR30 cell line by curcumin].

OBJECTIVE: To explore the effects of curcumin on paclitaxel resistance reversal of SKOV3-TR30 cell line and its mechanism. METHODS: The (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) MTT assay was performed to determine the sensitivity of curcumin-treated SKOV3-TR30 cells to paclitaxel. The cell cycle distribution of SKOV3-TR30 was analyzed by flow cytometry. And the expression level of glycogen synthase kinase-3 (GSK-3) protein was detected by Western blot. RESULTS: IC(50) of paclitaxel in SKOV3-TR30 decreased with a treatment of curcumin. And the reversal times was 3.0. Flow cytometric analysis of curcumin-treated SKOV3-TR30 cells demonstrated a distinct G(2)-M phase block (78.5 ± 6.4)% after a 12-hour treatment of paclitaxel versus SKOV3-TR30 cells without curcumin. There was a lack of G(2)-M phase arrest (only 27.0% ± 2.9%). The expression of GSK-3 protein in SKOV3-TR30 cells decreased with the 12 and 24-hour treatments of curcumin. CONCLUSION: Curcumin can partially reverse the paclitaxel-resistance of SKOV3-TR30 cells through a down-regulation of GSK-3.

A reversible S-adenosyl-L-homocysteine hydrolase inhibitor ameliorates experimental autoimmune encephalomyelitis by inhibiting T cell activation.

The reversible S-adenosyl-l-homocysteine hydrolase inhibitor DZ2002 [methyl 4-(adenin-9-yl)-2-hydroxybutanoate] suppresses antigen-induced-specific immune responses, particularly type 1 helper T cell (Th1)-type responses. Experimental autoimmune encephalomyelitis (EAE) is thought to be a Th1 cell-mediated inflammatory demyelinating autoimmune disease model of human multiple sclerosis (MS). In this study, we examined the effects of DZ2002 on active EAE induced by myelin oligodendrocyte glycoprotein (MOG) 35-55 in female C57BL/6 mice. Administration of DZ2002 (50 mg/kg/day i.p.) significantly reduced the incidence and severity of EAE, which was associated with the inhibition of MOG35-55-specific T cell proliferation and Th1-type cytokine production. In vitro studies also demonstrated that DZ2002 inhibited anti-CD3/28-induced naive T cell activation concomitant with the down-regulation of cyclin-dependent kinase (CDK) 4, CDK6, cyclin D3, and the up-regulation or protection of the CDK inhibitor p27. These findings highlight the fact that DZ2002 likely prevents EAE by suppressing T cell activation and suggest its utility in the treatment of MS and other Th1-mediated inflammatory diseases.

Differential expression of inducible nitric oxide synthase and IL-12 between peritoneal and splenic macrophages stimulated with LPS plus IFN-gamma is associated with the activation of extracellular signal-related kinase.

Resident peritoneal macrophages (pMphi) are found deficient in T cell-stimulating capacity compared with the competent splenic macrophages (sMphi). Macrophages (Mphi)-derived nitric oxide (NO) and IL-12 have been shown to play crucial roles in the interaction between Mphi and T cells. To further understand differential functions between pMphi and sMphi, we focused on the production of NO and IL-12 from LPS plus IFN-gamma-activated Mphi. We demonstrated the differential expression of inducible nitric oxide synthase (iNOS) and IL-12 in pMphi and sMphi with LPS plus IFN-gamma stimulation. pMphi produced high level of NO but low level of IL-12, whereas sMphi produced high level of IL-12 but no NO. Furthermore, we demonstrated that there were no differences in IFN-gamma-induced signal transducer and activator of transcription-1 activation and consequent interferon regulatory factor-1 and interferon consensus sequence-binding protein up-regulation between pMphi and sMphi. Likewise, p38 mitogen-activated protein kinase was activated by LPS with identical kinetics in both pMphi and sMphi. However, LPS-induced extracellular signal-regulated kinase (ERK) activation was prolonged in pMphi comparing with sMphi. Moreover, we demonstrated, using inhibitor selective for ERK cascade (PD98059), that the prolonged ERK activation contributed a positive signal for iNOS expression and a negative signal for IL-12p40 expression in resident pMphi. In addition, anti-IL-10-neutralizing antibody plus indomethacin could abrogate the inhibitory effects of endogenous IL-10 and prostaglandin E2 on the production of IL-12 by resident pMphi possibly through suppressing ERK activation. Taken together, profound difference in ERK activation may account for differential LPS plus IFN-gamma responsiveness between pMphi and sMphi. High production of NO and low production of IL-12 by pMphi may contribute to its deficiency in T cell-stimulating capacity.

S-adenosyl-L-homocysteine hydrolase inactivation curtails ovalbumin-induced immune responses.

The reversible S-adenosyl-L-homocysteine (AdoHcy) hydrolase inhibitor methyl 4-(adenin-9-yl)-2-hydroxybutanoate (DZ2002) suppresses macrophage activation and function. The effects of DZ2002 on T cell function, however, are still unclear. Here, we examined whether DZ2002 alters type 1 helper T cell (Th1) and/or type 2 helper T cell (Th2) immune responses, and whether these effects are associated with both the inhibition of AdoHcy hydrolase and intracellular elevation of endogenous AdoHcy. Male C57BL/6 mice immunized with ovalbumin (OVA) were treated with DZ2002 (1, 5, and 25 mg/kg/day) after which lymphocyte proliferation, cytokine production, and IgG responses to OVA were monitored. Administration of DZ2002 dose dependently suppressed OVA-specific lymphocyte proliferation and anti-OVA IgG production compared with controls. Interleukin (IL)-2 and interferon (IFN)-gamma as well as anti-OVA IgG2a and IgG3, indicators of Th1 immune responses, were markedly decreased in mice treated with DZ2002, whereas IL-4 and anti-OVA IgG1, indicators of Th2 immune responses, were only mildly suppressed. AdoHcy hydrolase activity in spleens of DZ2002-treated mice was substantially blocked, and not surprisingly, AdoHcy levels were significantly elevated compared with controls. Finally, similar immunosuppressive effects were also observed in mice treated with AdoHcy. These data strongly indicate that DZ2002 suppresses antigen-induced specific immune responses, particularly Th1 responses, through inhibition of AdoHcy hydrolase and elevation of endogenous AdoHcy.

Inhibition of S-adenosyl-L-homocysteine hydrolase induces immunosuppression.

Lymphocytes depend on transmethylation reactions for efficient activation and function. These reactions are primarily catalyzed by S-adenosylmethionine-dependent methyltransferases, which convert S-adenosylmethionine to S-adenosyl-L-homocysteine. S-adenosyl-L-homocysteine is then hydrolyzed by S-adenosyl-L-homocysteine hydrolase to prevent feedback inhibition of transmethylation reactions. By impeding S-adenosyl-L-homocysteine hydrolase, a build-up of S-adenosyl-L-homocysteine occurs, and most intracellular transmethylation reactions cease. Thus, a nontoxic inhibitor of this enzyme might be a useful immunosuppressive therapeutic agent. We identified a potent reversible type III inhibitor of S-adenosyl-L-homocysteine hydrolase, DZ2002 [methyl 4-(adenin-9-yl)-2-hydroxybutanoate], and determined its cytotoxic and immunologic effects. We demonstrated that DZ2002 blocked S-adenosyl-L-homocysteine hydrolase more effectively than a type I inhibitor, but cytotoxicity from DZ2002 was greatly reduced. Although DZ2002 did not prevent concanavalin A-induced T cell proliferation or interleukin (IL)-2 production, it significantly reduced both a mixed lymphocyte reaction and IL-12 production from in vitro-stimulated splenocytes. In addition, levels of CD80 and CD86 on human monocytic THP-1 cells were decreased in a dose-dependent manner in the presence of 0.1 to 10 microM DZ2002, and decreases were also seen in IL-12 and tumor necrosis factor-alpha production from both mouse thioglycollate-stimulated peritoneal macrophages and THP-1 cells. In vivo, DZ2002 significantly suppressed a delayed-type hypersensitivity reaction as well as antibody secretion. We conclude that DZ2002's immunosuppressive effects are likely not solely attributed to T cell inhibition but also to the obstruction of macrophage activation and function through reductions in cytokine output and/or T cell costimulation. These data suggest an important dual role for the S-adenosyl-l-homocysteine hydrolase in both macrophage and T cell function.

Effect of interleukin-7 gene transfection into ovarian carcinoma cell line SKOV3 in vitro and in vivo.

OBJECTIVE: To investigate the effect of interleukin-7 (IL-7) gene transfection into an established ovarian carcinoma cell line (SKOV3) in vitro and evaluate the tumorigenicity of SKOV3-IL-7 in severe combined immunodeficient (SCID) mice. METHODS: IL-7 gene was transfected into SKOV3 cells by liposome. IL-7 mRNA and protein of SKOV3-IL-7 and their parental control cells were detected by reverse transcriptive-polymerase chain reaction (RT-PCR) and Western blot, respectively. The levels of IL-7, IL-2, TNFalpha and TGFbeta1 in the supernatant were detected by ELISA. The cell cycle, HLA-ABC, HLA-DR and ICAM-1 expressions were assayed by flow cytometry. The sensitivities of tumor cells to lymphokine-activated killer (LAK) cells were measured by lactate dehydrogenase (LDH) release assay. Tumorigenicities of SKOV3-IL-7 and their parental cells in SCID mice were evaluated by macro- and histological examination, while IL-7 expression and secretion were detected by immunohistochemistry and ELISA. RESULTS: IL-7 mRNA and protein were detectable in SKOV3-IL-7 only. ICAM-1 expression was significantly higher and TGFbeta1 level in culture supernatants was significantly decreased in SKOV-IL-7, all other variables remained unchanged. The proliferative activity and cell cycle of SKOV3-IL-7 were unchanged. The cytotoxic sensitivity of SKOV3-IL-7 to LAK cells was significantly higher. Both gene-transfected and nontransfected SKOV3 cells were successfully inoculated into the peritoneal cavities of SCID mice. IL-7 proteins in plasmas and tumor tissues were detectable only in SCID mice inoculated with SKOV3-IL-7. The IL-7 engineered murine tumor models revealed better general aspects, reduced tumor development and dissemination. CONCLUSIONS: IL-7 gene transfection into SKOV3 cells downregulates TGFbeta1 secretion, upregulates ICAM-1 expression and enhances sensitivity to LAK cells in vitro. The tumorigenicity of IL-7 engineered cells in SCID mice is reduced. These findings may offer support to the development of cytokine gene therapy for ovarian carcinoma.

[Expression of mRNA isoforms of vascular endothelial growth factor in ovarian carcinoma]

OBJECTIVE: To study the expression of vascular endothelial growth factor (VEGF) mRNA isoforms in ovarian carcinoma and to explore their role in tumorigenesis and development of ovarian carcinoma. METHODS: The types and levels of VEGF mRNA isoforms of surgical samples from 30 patients with ovarian carcinoma were determined by relatively quantative RT-PCR, nest PCR and sequence analysis. RESULTS: VEGF(121), VEGF(145), VEGF(165) and VEGF(189)mRNA were detected in normal ovaries and ovarian carcinoma tissues. The expression level of VEGF(121) was significantly higher than that of VEGF(145), VEGF(165) and VEGF(189) (P<0.001, respectively). The expression of all 4 isoforms in carcinoma tissues was increased significantly compared with that in normal ovaries (P<0.05). CONCLUSION: Overexpression of VEGF(121), VEGF(145), VEGF (165) and VEGF(189) mRNA, especially VEGF(121), was found in varian carcinoma tissues. This findings suggest that all 4 VEGF isoforms may be involved in the tumorigenesis and development of ovarian carcinoma and VEGF(121) may play a key role.

The value of curettage in diagnosis of endometrial hyperplasia.

OBJECTIVES: The aim of this study was to assess the value of diagnosis of endometrial hyperplasia by curettage and to determine the results of proliferating cell nuclear antigen (PCNA) immunostaining in differentiating endometrial carcinoma from endometrial hyperplasia. METHODS: According to Kurman's criteria, we treated 150 patients with endometrial hyperplasia detected by curettage and compared retrospectively the diagnosis by curettage with that by hysterectomy. PCNA expression was examined using immunohistochemostaining on 60 patients with complex atypical hyperplasia detected by curettage. RESULTS: Simple hyperplasia was found by curettage in 53 patients, complex hyperplasia in 11, simple atypical hyperplasia in 26, and complex atypical hyperplasia in 60. All patients were rediagnosed after hysterectomy. As a result, 65 were found to have simple hyperplasia, 7 complex hyperplasia, 15 simple atypical hyperplasia, 29 complex atypical hyperplasia, and 34 endometrial carcinoma. The accuracy of histological diagnosis by curettage was 76.7-92.0% and was dependent on different types of hyperplasia. Simple atypical hyperplasia and complex atypical hyperplasia were more likely to coexist with endometrial carcinoma than both simple hyperplasia and complex hyperplasia (chi2 = 26.3, P < 0.001), and complex atypical hyperplasia was more likely to coexist with endometrial carcinoma than simple atypical hyperplasia (chi2 = 9.78, P < 0.005). In complex atypical hyperplasia patients, coexistence with endometrial carcinoma was more common after menopause than before menopause (chi2 = 3.93, P < 0.05). In complex atypical hyperplasia patients, the expression of PCNA in cases associated with endometrial carcinoma was higher or stronger than in cases associated without endometrial carcinoma (chi2 = 7.68, P < 0.01, or U = 252.00, P < 0.01). Conclusions. Curettage tends to be more highly accurate in diagnosing simple hyperplasia than complex atypical hyperplasia, which is often found by hysterectomy to be associated with endometrial carcinoma. The expression of PCNA may be helpful in differentiating complex atypical hyperplasia from endometrial carcinoma.