Elevated Intracranial Pressure Level Is a Risk Factor for Sepsis-associated Encephalopathy: A Prospective Cohort Study.

BACKGROUND/AIM: Cerebral edema is common in patients with sepsis-associated encephalopathy (SAE) and is a major cause of elevated intracranial pressure (ICP); however, the relationship between elevated ICP and SAE is unclear. The aim of this study was to investigate the association between optic nerve sheath diameter (ONSD), a surrogate of ICP, and the incidence of SAE. PATIENTS AND METHODS: A prospective observational study was performed in a medical-surgical adult intensive care unit (ICU). All patients in the ICU who were consecutively diagnosed with sepsis during the study period were evaluated for eligibility. Ultrasound measurements of ONSD were performed within 6 h of enrollment and every two days thereafter until the patient developed SAE. Clinical and blood test data were collected throughout this period. Patients underwent a daily conscious and cognitive assessment. SAE was diagnosed as delirium or Glasgow Coma Scale (GCS) <15 points. Multivariate modified Poisson regression analysis was performed to identify risk factors for SAE. RESULTS: A total of 123 patients with sepsis were included in the analysis. 58 patients (47.2%) developed SAE. The levels of ONSD0 (the first measured value) and ONSDmax (the maximum measured value) in the SAE group were significantly higher than those in the non-SAE group (5.23±0.52 mm vs. 5.85±0.54 mm for ONSD0 and 5.41±0.46 mm vs. 6.09±0.58 mm for ONSDmax, respectively; all p-values <0.001). The area under the curves (AUCs) for the ONSD0 and ONSDmax values in predicting SAE were 0.801 (95%CI=0.723-0.880, p<0.001) and 0.829 (95%CI=0.754-0.903, p<0.001), respectively. A higher ONSD0 level was significantly associated with an increased risk of SAE (adjusted risk ratio 3.241; 95%CI=1.686-6.230, p<0.001). CONCLUSION: The levels of ONSD correlate with risk of SAE, indicating that increased ICP level is an independent risk factor for the development of SAE. Dynamic monitoring of ONSD/ICP has a high predictive value for SAE. Measures to prevent increases in ICP are helpful to reduce the incidence of SAE in sepsis patients.

Research Progress on the Microregulatory Mechanisms of Fertilization: A Review.

The process of fertilization includes sperm capacitation, hyperactivation, an acrosome reaction and the release of acrosome enzymes, membrane fusion and channel formation, the release of the sperm nucleus, and gamete fusion. This process is closely related to the shape and vitality of the sperm, acrosome enzyme release, and the zona pellucida structure of the egg, as well as the opening and closing of various ion (e.g., calcium) channels, the regulation of signaling pathways such as cyclic adenosine monophosphate-protein kinase A, the release of progesterone, and the coupling of G-proteins. The interaction among multiple factors and their precise regulation give rise to multiple cascading regulatory processes. Problems with any factor will affect the success rate of fertilization. Recent studies have shown that with rapid societal development, the incidence of male infertility is increasing and occurs at younger ages. According to World Health Organization statistics, 15% of couples of childbearing ages have infertility problems, of which 50% are caused by male factors. Additionally, the cause of infertility cannot be identified in as many as 60% to 75% of male infertility patients. In this article, we review the research progress on the microregulation of fertilization and mechanisms underlying this process to identify causes and develop novel prevention and treatment strategies for male infertility.

Diabetes Mellitus Causes Male Reproductive Dysfunction: A Review of the Evidence and Mechanisms.

The metabolic disorders caused by diabetes can lead to various complications, including dysfunction of the male reproductive system. In patients with diabetes, long-term hyperglycemia results in diabetic vascular neuropathy, oxidative stress injury, abnormal zinc metabolism, and insulin resistance syndrome. In addition, insulin deficiency and resistance in diabetes can damage the hypothalamus, pituitary gland, gonads, and perigonads. This can reduce the secretion of sex hormones including gonadotropin-releasing hormone, follicle stimulating hormone, luteinizing hormone, and testosterone, and can lead to testicular atrophy, stromal cell atrophy, seminiferous tubule damage, spermatogenic cell damage, and other structural injuries of the male reproductive organs. These actions can affect male fertility and reproductive health. Herein, we review studies that report a causative role of diabetes in male reproductive function. We also discuss the evidence-based mechanisms involved in the processes of diabetes-related male sexual and reproductive dysfunction as well as the progress in treatment.

Finite Element Analysis of an Improved Correction System for Spinal Deformity.

BACKGROUND/AIM: Surgical treatment for spinal deformity aims to correct malformation, release the nerves, and reconstruct spinal stability. To explore and develop a new improved spinal correction system (ISCS) for clinical application, we studied the stability and biomechanical characteristics of the ISCS through finite element analysis and comparison of the ISCS with the pedicle screw and rod system (PSRS). PATIENTS AND METHODS: Using L1-L3 CT image data of a normal adult male lumbar spine for establishment of L1-L3 finite element model, we established posterior internal fixation models for a comparative finite element analysis of PSRS and ISCS. An axial load of 500 N and a moment of 10 N•m were applied to L1 to simulate flexion, extension, lateral bending, and axial rotation. Stress distribution characteristics, load sharing, strain bending stiffness and strain angle change of the models were measured. RESULTS: In flection and extension directions, the maximum stress of the L2 vertebral body and the L1/2 and L2/3 discs in PSRS was less than that of ISCS. In lateral bending and axial rotation directions, the maximum stress between PSRS and ISCS was similar. However, the stress shielding rate of L2, L1/2, and L2/3 intervertebral discs in ISCS was significantly lower than that of PSRS. We also found that both models had similar angular displacement and maximum displacement in lateral bending direction, but PSRS had a lower angular displacement and maximum displacement in flection and extension directions. Finally, we showed that PSRS had similar angular displacement and a lower maximum displacement compared with ISCS in axial rotation, whereas ISCS had lower bending stiffness than PSRS in different directions. CONCLUSION: ISCS can effectively fix spinal deformities compared to PSRS. ISCS provides a new option for orthopedic surgery treatment of scoliosis and, therefore, warrants further clinical studies in patients with other spinal deformities.

High Rate of Detection of Human ESPL1-HBV S Fusion Gene in Patients With HBV-related Liver Cancer: A Chinese Case-Control Study.

AIM: It has been shown that the integration of hepatitis B virus (HBV) gene into the host genome is a high-risk factor for development of hepatocellular carcinoma (HCC). However, the relationship between HBV S-integrated human extra spindle pole bodies-like 1 (ESPL1) gene and HCC is unknown. This study was designed to detect HBV S-integrated human ESPL1 fusion gene in patients with HCC for potentially using this fusion gene as a biomarker for HCC diagnosis. PATIENTS AND METHODS: Nineteen and 70 patients with chronic hepatitis B (CHB) were recruited to the experimental and control groups, respectively, and both groups underwent an effective nucleoside/nucleotide analog therapy and follow-up for HCC occurrence for up to 11 years. HCC tissues were obtained by surgical resection from the experimental group, while liver tissues were collected by liver biopsy in the control group prior to treatment with nucleoside/nucleotide analogs. Alu polymerase chain reaction was used to assess HBV S gene integration in the liver tissues from both groups. HBV S-integrated human ESPL1 fusion gene was then detected in patients with HBV S gene integration using a gene database. RESULTS: All patients in the experimental group developed HCC, whereas no HCC was diagnosed in the control group. HBV S gene integration was identified in 12 out of 19 HCC tissues in the experimental group, giving a detection rate of 63.2%, which was significantly greater than that of 15.7% (11/70) in the control group (p<0.001). We further showed that HBV S-integrated human ESPL1 fusion gene was detected in eight patients (rate of 66.7%) among the 12 patients with HCC with HBV S gene integration in the experimental group, whereas the fusion gene was not detectable in any of the patients in the control group (p=0.001). CONCLUSION: This research demonstrates a high detection rate of HBV S-integrated human ESPL1 fusion gene in patients with HBV-related HCC and shows that this fusion gene appears to be associated with HCC development in patients with CHB. These findings suggest that HBV S-integrated human ESPL1 fusion gene may potentially serve as a biomarker for early detection of HCC in HBV-infected populations.

Sterol uptake and sterol biosynthesis act coordinately to mediate antifungal resistance in Candida glabrata under azole and hypoxic stress.

Pathogenic fungi, including Candida glabrata, develop strategies to grow and survive both in vitro and in vivo under azole stress. However, the mechanisms by which yeast cells counteract the inhibitory effects of azoles are not completely understood. In the current study, it was demonstrated that the expression of the ergosterol biosynthetic genes ERG2, ERG3, ERG4, ERG10, and ERG11 was significantly upregulated in C. glabrata following fluconazole treatment. Inhibiting ergosterol biosynthesis using fluconazole also increased the expression of the sterol influx transporter AUS1 and the sterol metabolism regulators SUT1 and UPC2 in fungal cells. The microarray study quantified 35 genes with elevated mRNA levels, including AUS1, TIR3, UPC2, and 8 ERG genes, in a C. glabrata mutant strain lacking ERG1, indicating that sterol importing activity is increased to compensate for defective sterol biosynthesis in cells. Bioinformatic analyses further revealed that those differentially expressed genes were involved in multiple cellular processes and biological functions, such as sterol biosynthesis, lipid localization, and sterol transport. Finally, to assess whether sterol uptake affects yeast susceptibility to azoles, we generated a C. glabrata aus1∆ mutant strain. It was shown that loss of Aus1p in C. glabrata sensitized the pathogen to azoles and enhanced the efficacy of drug exposure under low oxygen tension. In contrast, the presence of exogenous cholesterol or ergosterol in medium rendered the C. glabrata AUS1 wild­type strain highly resistant to fluconazole and voriconazole, suggesting that the sterol importing mechanism is augmented when ergosterol biosynthesis is suppressed in the cell, thus allowing C. glabrata to survive under azole pressure. On the basis of these results, it was concluded that sterol uptake and sterol biosynthesis may act coordinately and collaboratively to sustain growth and to mediate antifungal resistance in C. glabrata through dynamic gene expression in response to azole stress and environmental challenges.

Protein kinase D inhibitor CRT0066101 suppresses bladder cancer growth in vitro and xenografts via blockade of the cell cycle at G2/M.

The protein kinase D (PKD) family of proteins are important regulators of tumor growth, development, and progression. CRT0066101, an inhibitor of PKD, has antitumor activity in multiple types of carcinomas. However, the effect and mechanism of CRT0066101 in bladder cancer are not understood. In the present study, we show that CRT0066101 suppressed the proliferation and migration of four bladder cancer cell lines in vitro. We also demonstrate that CRT0066101 blocked tumor growth in a mouse flank xenograft model of bladder cancer. To further assess the role of PKD in bladder carcinoma, we examined the three PKD isoforms and found that PKD2 was highly expressed in eight bladder cancer cell lines and in urothelial carcinoma tissues from the TCGA database, and that short hairpin RNA (shRNA)-mediated knockdown of PKD2 dramatically reduced bladder cancer growth and invasion in vitro and in vivo, suggesting that the effect of the compound in bladder cancer is mediated through inhibition of PKD2. This notion was corroborated by demonstrating that the levels of phospho-PKD2 were markedly decreased in CRT0066101-treated bladder tumor explants. Furthermore, our cell cycle analysis by flow cytometry revealed that CRT0066101 treatment or PKD2 silencing arrested bladder cancer cells at the G2/M phase, the arrest being accompanied by decreases in the levels of cyclin B1, CDK1 and phospho-CDK1 (Thr161) and increases in the levels of p27Kip1 and phospho-CDK1 (Thr14/Tyr15). Moreover, CRT0066101 downregulated the expression of Cdc25C, which dephosphorylates/activates CDK1, but enhanced the activity of the checkpoint kinase Chk1, which inhibits CDK1 by phosphorylating/inactivating Cdc25C. Finally, CRT0066101 was found to elevate the levels of Myt1, Wee1, phospho-Cdc25C (Ser216), Gadd45α, and 14-3-3 proteins, all of which reduce the CDK1-cyclin B1 complex activity. These novel findings suggest that CRT0066101 suppresses bladder cancer growth by inhibiting PKD2 through induction of G2/M cell cycle arrest, leading to the blockade of cell cycle progression.

Microsatellite analysis of genetic diversity in the Tupaia belangeri yaoshanensis.

The Chinese tree shrew (Tupaia belangeri yaoshanensis) has long been proposed to serve as an animal model for studying human diseases. However, its overall genetic diversity and population structure remain largely unknown. In the present study, we investigated the genetic diversity of population microsatellite DNA in wild Tupaia belangeri yaoshanensis. Sixteen microsatellite loci were assessed in 76 wild Tupaia belangeri yaoshanensis. The target microsatellite DNA fragments were amplified from the peripheral blood DNA of the animals by polymerase chain reaction (PCR), and the PCR-amplified products were verified by DNA sequencing and used for the analysis of allele, effective allele, genetic heterozygosity, polymorphism and population structure. Our results showed that of the 16 microsatellite loci examined, 5 microsatellite loci were monomorphic and 11 microsatellite loci were polymorphic. We detected 61 alleles in the polymorphic loci and found 2-10 (with an average of 5.5455) alleles per locus. Our data also showed that the observed and expected heterozygosities ranged from 0.087 to 0.8947 and 0.1368 to 0.7892 with an average of 0.3968 and 0.4796, respectively. Taken together, the results revealed a considerably high heterozygosity and high genetic diversity at the molecular level in the population of wild Tupaia belangeri yaoshanensis. The identified markers from the present study may be useful for individual identification and parentage testing, as well as for the quantification of population heterogeneity in the Chinese tree shrew.

Proteomic analysis of proteome and histone post-translational modifications in heat shock protein 90 inhibition-mediated bladder cancer therapeutics.

Heat shock protein 90 (HSP90) inhibition is an attractive strategy for cancer treatment. Several HSP90 inhibitors have shown promising effects in clinical oncology trials. However, little is known about HSP90 inhibition-mediated bladder cancer therapy. Here, we report a quantitative proteomic study that evaluates alterations in protein expression and histone post-translational modifications (PTMs) in bladder carcinoma in response to HSP90 inhibition. We show that 5 HSP90 inhibitors (AUY922, ganetespib, SNX2112, AT13387, and CUDC305) potently inhibited the proliferation of bladder cancer 5637 cells in a dose- and time-dependent manner. Our proteomic study quantified 518 twofold up-regulated and 811 twofold down-regulated proteins common to both AUY922 and ganetespib treatment. Bioinformatic analyses revealed that those differentially expressed proteins were involved in multiple cellular processes and enzyme-regulated signaling pathways, including chromatin modifications and cell death-associated pathways. Furthermore, quantitative proteome studies identified 14 types of PTMs with 93 marks on the core histones, including 34 novel histone marks of butyrylation, citrullination, 2-hydroxyisobutyrylation, methylation, O-GlcNAcylation, propionylation, and succinylation in AUY922- and ganetespib-treated 5637 cells. Together, this study outlines the association between proteomic changes and histone PTMs in response to HSP90 inhibitor treatment in bladder carcinoma cells, and thus intensifies the understanding of HSP90 inhibition-mediated bladder cancer therapeutics.

Histone deacetylase inhibitor-induced cell death in bladder cancer is associated with chromatin modification and modifying protein expression: A proteomic approach.

The Cancer Genome Atlas (TCGA) project recently identified the importance of mutations in chromatin remodeling genes in human carcinomas. These findings imply that epigenetic modulators might have a therapeutic role in urothelial cancers. To exploit histone deacetylases (HDACs) as targets for cancer therapy, we investigated the HDAC inhibitors (HDACIs) romidepsin, trichostatin A, and vorinostat as potential chemotherapeutic agents for bladder cancer. We demonstrate that the three HDACIs suppressed cell growth and induced cell death in the bladder cancer cell line 5637. To identify potential mechanisms associated with the anti-proliferative and cytotoxic effects of the HDACIs, we used quantitative proteomics to determine the proteins potentially involved in these processes. Our proteome studies identified a total of 6003 unique proteins. Of these, 2472 proteins were upregulated and 2049 proteins were downregulated in response to HDACI exposure compared to the untreated controls (P<0.05). Bioinformatic analysis further revealed that those differentially expressed proteins were involved in multiple biological functions and enzyme-regulated pathways, including cell cycle progression, apoptosis, autophagy, free radical generation and DNA damage repair. HDACIs also altered the acetylation status of histones and non-histone proteins, as well as the levels of chromatin modification proteins, suggesting that HDACIs exert multiple cytotoxic actions in bladder cancer cells by inhibiting HDAC activity or altering the structure of chromatin. We conclude that HDACIs are effective in the inhibition of cell proliferation and the induction of apoptosis in the 5637 bladder cancer cells through multiple cell death-associated pathways. These observations support the notion that HDACIs provide new therapeutic options for bladder cancer treatment and thus warrant further preclinical exploration.

Taurine Attenuates Dimethylbenz[a]anthracene-induced Breast Tumorigenesis in Rats: A Plasma Metabolomic Study.

Breast cancer is the most common malignancy and the leading cause of cancer-related mortality in women worldwide. Taurine, the most abundant free amino acid, plays a role in several biological processes in humans and has been shown to have activity against breast cancer and other tumors. To investigate the role and mechanism of taurine action in breast cancer, we used dimethylbenz[a]anthracene (DMBA)-induced breast carcinogenesis in rats as a model of breast cancer. The administration of taurine significantly reduced the DMBA-induced breast cancer rate from 80% to 40% in rats (p<0.05). Metabolomic studies using time-of-flight gas chromatography-mass spectrometry identified 23 differential metabolites in the plasma of taurine-administered rats. Bioinformatic analysis further revealed that these metabolites are involved in multiple metabolic pathways, including energy, glucose, amino acid, and nucleic acid metabolism, suggesting that the antitumor activity of taurine in rats is mediated through altered metabolism of breast cancer cells. We propose that these differential metabolites may be potential biomarkers for monitoring cancer therapy and prognosis in the clinic. This study provides a scientific basis for further investigations of the antitumor mechanism of taurine and the development of novel therapeutic strategies to treat breast cancer.

Perfusion of a cerebral protective solution enhances neuroprotection in a rabbit model of occlusion-reperfusion: prolonged cerebral dormancy time.

In the present study, we investigated the effect of a cerebral protective solution on prolongation of cerebral dormancy time in a rabbit model of occlusion-reperfusion. In a control group, rabbits were anesthetized and the four cerebral arteries (the left and right common carotid arteries and vertebral arteries) were occluded for 7.5 min followed by reperfusion. All six rabbits in the control group died. In contrast, a second group underwent perfusion of a cerebral protective solution for 15 min between artery occlusion and reperfusion. All six rabbits in this group survived. However, when the perfusion solution was changed to 5% glucose solution or rabbit plasma in two other groups, the rabbits in both the latter two groups also died. Neuroprotection was also observed when the protective solution was administered for 30-60 min after the onset of artery occlusion and before the return of blood flow (reperfusion). To understand the high rate of thrombotic stroke in the clinic, we assessed the influence of different organ tissue infusions on blood coagulation in vitro and found that blood clotting occurred faster in the presence of brain tissue infusion compared to liver, kidney, and heart tissue infusions. These results indicate a higher rate of thrombosis in brain tissue compared to any of the other tissues tested. The current study shows that perfusion of a cerebral protective solution produced a significant neuroprotective benefit in our rabbit model of occlusion-reperfusion, suggesting that administration of a cerebral protective solution may be an effective approach for the treatment of ischemic stroke.

Advances in intravesical therapy for the treatment of non-muscle invasive bladder cancer (Review).

The knowledge of tumor biology and the biomechanical properties of the urothelium have led to significant advances in the development of intravesical therapy for the treatment of non-muscle invasive bladder cancer (NMIBC). Targeted therapy improves the efficacy and decreases the side effects of antineoplastic agents. Nanoparticles that target antitumor agents to the urothelial cells have allowed for improved delivery of these agents to tumor cells. Gene therapy is another strategy that has allowed for a targeted induction of an antitumor response. Finally, engineering of the bacillus Calmette-Guérin (BCG) vaccine aimed to minimize the potential side effects associated with this treatment. These novel approaches hold promise for decreasing the rate of progression and recurrence of NMIBC.

Selective tumor cell killing by triptolide in p53 wild-type and p53 mutant ovarian carcinomas.

Triptolide is a traditional Chinese medicinal herb-derived antineoplastic agent. However, its antitumor activity against gynecologic carcinomas has not yet been well described. It is the purpose of this article to investigate the effect and mechanism of triptolide in human ovarian cancer using both A2780 (p53 wild) and OVCAR-3 (p53 mutated) cells. Our results showed that triptolide exerted a potent inhibitory effect on the growth and proliferation of both cell lines in a dose- and time-dependent manner and that the effect was independent of the expression of p53. In contrast, triptolide had only a marginal cytotoxicity in noncancerous ovary cells, lung fibroblast cells, and macrophage cells, indicating differential inhibitory effects of the drug on cell growth between ovarian cancer cells and normal tissue cells. Exposure of the ovarian cancer cells to triptolide induced apoptosis, as evaluated by annexin V/propidium iodide-labeled flow cytometry. Triptolide-induced apoptosis was accompanied by cytochrome c release and caspase-3 activation and was associated with downregulation of Bcl-2 and upregulation of Bax. Cell cycle analysis demonstrated that treatment with triptolide induced cell cycle S phase arrest in A2780 cells and G2/M phase arrest in OVCAR-3 cells. Further detection by Western blotting revealed that the cell cycle arrest by triptolide in both cell lines occurred in concert with increased expression of p21(CIP1/WAF1). This study shows that triptolide selectively kills ovarian cancer cells with different p53 status predominantly through regulating the coordinate and dynamic cellular processes of proliferation and apoptosis, thereby making it a promising chemotherapeutic agent against a broad spectrum of ovarian carcinomas.

A novel approach for enriching cancer stem cells from the human SW-13 adrenocortical carcinoma cell line.

The present study was undertaken to develop a new method for enriching cancer stem cells (CSCs) from the human adrenal cortical carcinoma (ACC) cell line SW-13. Given that the existence of CSCs in ACC causes resistance to conventional chemotherapies, treatment with cyclophosphamide was used for in vivo selection of CSCs in a BALB/c nude mouse tumor xenograft model established using the ACC cell line SW-13. The characteristics of CSCs in three generations of tumor xenografts were assessed for single-cell colony formation, flat colony formation, and cell sphere formation in serum-free suspension culture. The formation rates of single-cell colonies, flat colonies, and cell spheres were significantly higher for tumor xenograft cells treated with cyclophosphamide than for untreated engrafted tumor cells. Flow cytometry to examine expression of the CSC markers C-X-C chemokine receptor type-4 (CXCR4; CD184) and ATP-binding cassette sub-family G member-2 (ABCG2; CDw338) revealed markedly higher levels of CXCR4 and ABCG2 in cyclophosphamide-treated xenograft tumor cells compared to untreated tumor cells. Together, these results indicate that cyclophosphamide treatment of tumor xenograft cells caused enrichment of CSCs with a strong capability for self-renewal and proliferation. In this method, the administration of cyclophosphamide selectively kills cancer cells without toxicity to CSCs and thereby provides a practical approach for achieving the enrichment of CSCs in ACC.

Enhancement of cisplatin-induced apoptosis by ß-elemene in resistant human ovarian cancer cells.

ß-Elemene is a new anticancer compound extracted from the Chinese medicinal herb Rhizoma zedoariae. We have shown previously that ß-elemene increases cisplatin cytotoxicity and enhances cisplatin sensitivity via blocking cell cycle progression at G2/M phase in resistant ovarian tumor cells. In the current study, we asked whether ß-elemene-augmented cisplatin activity in ovarian carcinoma cells is mediated through the induction of apoptosis. Here, we show that ß-elemene triggered apoptotic cell death in chemoresistant human ovarian cancer A2780/CP and MCAS cells in a dose- and time-dependent fashion, as assessed by six different apoptosis assays. Intriguingly, ß-elemene was a stronger inducer of apoptosis than cisplatin in this model system, and a synergistic effect on induction of cell death was observed when the tumor cells were treated with both agents. Furthermore, ß-elemene plus cisplatin exposure significantly disrupted the mitochondrial transmembrane potential (ΔΨ (m)) and increased the release of cytochrome c from mitochondria into the cytoplasm. The combination treatment with both compounds also induced increases in caspase-3/8/9 activities and caspase-9 cleavage, enhanced protein expression of Bax and phosphorylation of Bcl-2 at Ser-70, and reduced the protein levels of Bcl-2 and Bcl-X(L) in the platinum-resistant ovarian cancer cells. Taken together, these data indicate that ß-elemene sensitizes chemoresistant ovarian carcinoma cells to cisplatin-induced apoptosis and that the augmented effect of ß-elemene on cisplatin cytotoxicity and sensitivity in resistant ovarian tumor cells is mediated through a mitochondria- and caspase-dependent cell death pathway.

Anticancer activity of ß-Elemene and its synthetic analogs in human malignant brain tumor cells.

Malignant brain tumors are aggressive in both children and adults. Despite recent improvements in diagnostic techniques, therapeutic approaches remain disappointing and unsuccessful. There is an urgent need for promising anticancer agents to improve overall survival of patients with brain cancer. ß-Elemene has been shown to have antiproliferative effects on many types of carcinomas. In this study, we compared the cytotoxic efficacy of ß-elemene and its synthetic analogs in the brain tumor cell lines A172, CCF-STTG1, and U-87MG. ß-Elemene exhibited cytotoxicity towards the tumor lines, effectively suppressing tumor cell survival. The inhibitory effect of ß-elemene was mediated by the induction of apoptosis, as demonstrated by three assays. The annexin V assay showed that ß-elemene increased the percentage of early- and late-apoptotic cells. Apoptotic nuclei were detected in cancer cells in situ by the terminal deoxynucleotidyltransferase-mediated deoxy-UTP-fluorescein nick end labeling (TUNEL) staining, and the number of TUNEL-positive cells was significantly increased at 24-72 h following drug treatment of the cell lines. Cell death enzyme-linked immunosorbent assay (ELISA) gave similar results. Furthermore, ß-elemene increased caspase-3/7/10 activity, up-regulated protein expression of BAX, and down-regulated the one of BCL-2, BCL-XL, and of X-linked inhibitor of apoptosis (XIAP) in the cells, suggesting that apoptotic signaling pathways are involved in the responses triggered by ß-elemene. Compared with ß-elemene, only three of the 10 synthetic ß-elemene analogs studied here, exerted comparable cytotoxic efficacy towards the three brain tumor lines: the analogs Lr-1 and Lr-2 had the same antitumor efficacy, while Lr-3 was less potent than ß-elemene. Thus, some synthetic analogs of ß-elemene may inhibit brain cancer cell growth and proliferation, and the synthetic analogs Lr-1 and Lr-2 may have great potential as alternatives to ß-elemene for anticancer therapy. Overall, this study provides, to our knowledge, the first evidence showing that synthetic analogs of ß-elemene hold promise for patients with brain tumors.

Syphilis manifesting as a nasopharyngeal carcinoma with cervical lymphadenopathy: A case report.

The present case report describes a case of syphilitic lymphadenopathy and raises the awareness of the differential diagnosis of cervical lymphadenopathy. A 50-year-old male worker presented with a 6-month history of enlarged and growing lymph nodes in the right upper neck and a blood-tinged post-nasal drip. Physical examination showed multiple enlarged lymph nodes located in the right upper neck. On nasopharyngoscopy, a mass was found in the nasopharynx. The histopathology of both the nasopharyngeal mass and the enlarged lymph nodes revealed non-specific inflammation. Rapid plasma reagin test results (titer, 1:1280) and Treponema pallidum particle assay results (titer, 1:2560) were positive. Subsequently, a diagnosis of syphilis was confirmed clinically and serologically. The reaction after penicillin treatment further confirmed the syphilis diagnosis. Thus, syphilis should be considered as a possibility in the differential diagnosis of cervical lymphadenopathy.

Case report of concurrent primary malignancies of the breast and nasopharynx.

The aim of this study was to report a case of concurrent primary malignancies of the breast and nasopharynx and discuss the potential relationship between Epstein-Barr virus (EBV) infection and breast cancer. A 39-year-old female presented with a palpable mass present for 1 year in her left breast. Immunohistochemical staining was performed and the results showed that the tumor cells were immunopositive for the estrogen receptor, progesterone receptor and p53 protein, and markedly positive for C-erb B2. In addition, 30% of the tumor cells were positive for the Ki-67 antigen. Blood test results revealed that EBV-CA-IgG was present and EBV-EA-IgG was reactivated. The patient was diagnosed with breast cancer (T1N0M0) and EBV infection. A mastectomy with axillary clearance was performed on the left breast. Histopathological examination provided evidence of invasive ductal adenocarcinoma. Further evaluation due to epistaxis following the breast surgery resulted in a diagnosis of nasopharyngeal carcinoma (T2N1M0). Histopathology showed a non-keratinizing undifferentiated carcinoma. The patient was treated with chemoradiotherapy for nasopharyngeal carcinoma. Twelve months following surgery and chemoradiotherapy the patient was assessed at the Cancer Hospital of Guangxi Medical University outpatient clinic and no evidence of relapse or metastasis was found. Thus, EBV infection may be involved in the pathogenesis of breast cancer, as observed in nasopharyngeal carcinoma.

Evaluation of reference genes for real-time quantitative PCR studies in Candida glabrata following azole treatment.

BACKGROUND: The selection of stable and suitable reference genes for real-time quantitative PCR (RT-qPCR) is a crucial prerequisite for reliable gene expression analysis under different experimental conditions. The present study aimed to identify reference genes as internal controls for gene expression studies by RT-qPCR in azole-stimulated Candida glabrata. RESULTS: The expression stability of 16 reference genes under fluconazole stress was evaluated using fold change and standard deviation computations with the hkgFinder tool. Our data revealed that the mRNA expression levels of three ribosomal RNAs (RDN5.8, RDN18, and RDN25) remained stable in response to fluconazole, while PGK1, UBC7, and UBC13 mRNAs showed only approximately 2.9-, 3.0-, and 2.5-fold induction by azole, respectively. By contrast, mRNA levels of the other 10 reference genes (ACT1, EF1α, GAPDH, PPIA, RPL2A, RPL10, RPL13A, SDHA, TUB1, and UBC4) were dramatically increased in C. glabrata following antifungal treatment, exhibiting changes ranging from 4.5- to 32.7-fold. We also assessed the expression stability of these reference genes using the 2(-ΔΔCT) method and three other software packages. The stability rankings of the reference genes by geNorm and the 2(-ΔΔCT) method were identical to those by hkgFinder, whereas the stability rankings by BestKeeper and NormFinder were notably different. We then validated the suitability of six candidate reference genes (ACT1, PGK1, RDN5.8, RDN18, UBC7, and UBC13) as internal controls for ten target genes in this system using the comparative CT method. Our validation experiments passed for all six reference genes analyzed except RDN18, where the amplification efficiency of RDN18 was different from that of the ten target genes. Finally, we demonstrated that the relative quantification of target gene expression varied according to the endogenous control used, highlighting the importance of the choice of internal controls in such experiments. CONCLUSIONS: We recommend the use of RDN5.8, UBC13, and PGK1 alone or the combination of RDN5.8 plus UBC13 or PGK1 as reference genes for RT-qPCR analysis of gene expression in C. glabrata following azole treatment. In contrast, we show that ACT1 and other commonly used reference genes (GAPDH, PPIA, RPL13A, TUB1, etc.) were not validated as good internal controls in the current model.