Exosomes based strategies for cardiovascular diseases: Opportunities and challenges.

Exosomes, as nanoscale extracellular vesicles (EVs), are secreted by all types of cells to facilitate intercellular communication in living organisms. After being taken up by neighboring or distant cells, exosomes can alter the expression levels of target genes in recipient cells and thereby affect their pathophysiological outcomes depending on payloads encapsulated therein. The functions and mechanisms of exosomes in cardiovascular diseases have attracted much attention in recent years and are thought to have cardioprotective and regenerative potential. This review summarizes the biogenesis and molecular contents of exosomes and details the roles played by exosomes released from various cells in the progression and recovery of cardiovascular disease. The review also discusses the current status of traditional exosomes in cardiovascular tissue engineering and regenerative medicine, pointing out several limitations in their application. It emphasizes that some of the existing emerging industrial or bioengineering technologies are promising to compensate for these shortcomings, and the combined application of exosomes and biomaterials provides an opportunity for mutual enhancement of their performance. The integration of exosome-based cell-free diagnostic and therapeutic options will contribute to the further development of cardiovascular regenerative medicine.

Intervention with ICOSL Antibodies Alleviates Inflammatory Infiltrations in Mice with Neutrophilic Asthma.

Background: Neutrophilic asthma is characterized by the predominant infiltration of neutrophils in airway inflammation. Objective: To explore the therapeutic potential of an antibody against the inducible T cell co-stimulator ligand (ICOSL) in a mouse model of neutrophilic asthma. Methods: Female BALB/c mice were randomly assigned to different groups. They were then injected with ovalbumin (OVA)/lipopolysaccharides (LPS) to induce neutrophilic asthma. The mice were then treated with either anti-ICOSL (the I group), control IgG (the G group), or no treatment (the N group). Additionally, a control group of mice received vehicle PBS and was labeled as the C group (n=6 per group). One day after the last allergen exposure, cytokine levels were measured in plasma and bronchoalveolar lavage fluid (BALF) using ELISA. After analyzing and categorizing BALF cells, the lung tissues were examined histologically and immunohistochemically. Results: Administering anti-ICOSL resulted in a significant decrease in the total number of inflammatory infiltrates and neutrophils found in BALF. Moreover, it led to a decrease in the levels of interleukin (IL)-6, IL-13, and IL-17 in both BALF and plasma. Additionally, there was an increase in IFN-γ levels in the BALF of asthmatic mice (p<0.05 for all). Treatment with anti-ICOSL also reduced lung interstitial inflammation, mucus secretion, and ICOSL expression in asthmatic mice. Conclusion: The treatment of anti-ICOSL effectively improved lung interstitial inflammation and mucus secretion in mice with neutrophilic asthma by restoring the balance of Th1/Th2/Th17 responses. These findings indicate that blocking the ICOS/ICOSL signaling could be an effective way to manage neutrophilic asthma.

Mitochondrial KATP channel-mediated autophagy contributes to angiotensin II-induced vascular dysfunction in mice.

BACKGROUND AND AIM: The present study aimed to investigate whether the mitochondrial KATP channel contributes to angiotensin II (Ang II)-induced vascular dysfunction, the development of hypertension, and atherosclerosis. METHODS AND RESULTS: ApoE (-/-) mice fed a high-fat diet were chronically infused with Ang II for eight weeks and concomitantly treated with losartan (ARB), apocynin, or 5-hydroxy decanoate (5-HD), or 3-methyladenine (3-MA). Systolic blood pressure was measured, and pathological changes of aortic or liver tissue were observed. Nitric oxide (NO), superoxide dismutase 2 (SOD2) levels and vasorelaxation rate were measured, and protein and mRNA expressions were examined by western blot and RT-PCR. Ang II-induced development of hypertension was suppressed not only by ARB, and apocynin but also by 5-HD or 3-MA. Ang II infusion decreased aortic NO production and relaxation, as well as SOD2 activity in liver, which were improved by all treatments. In addition, Ang II-induced activation of autophagy was suppressed by 5-HD in aortic tissue, furthermore, Ang II increases the atherosclerotic index in plasma and exacerbates the development of atherosclerosis by increases of fat deposition in the aorta and liver. Lipid metabolism-related mRNA expressions (LXR-α, LDLR, SRBI, Acca, and FASN) were changed by Ang II. Similarly, not only ARB, and apocynin, but also 5-HD and 3-MA suppressed Ang II-induced these changes. CONCLUSIONS: Our present findings evidence that mitochondrial KATP channel-mediated autophagy contributes to Ang II-induced vascular dysfunction, development of hypertension, and atherosclerosis.

Comprehensive view of macrophage autophagy and its application in cardiovascular diseases.

Cardiovascular diseases (CVDs) are the primary drivers of the growing public health epidemic and the leading cause of premature mortality and economic burden worldwide. With decades of research, CVDs have been proven to be associated with the dysregulation of the inflammatory response, with macrophages playing imperative roles in influencing the prognosis of CVDs. Autophagy is a conserved pathway that maintains cellular functions. Emerging evidence has revealed an intrinsic connection between autophagy and macrophage functions. This review focuses on the role and underlying mechanisms of autophagy-mediated regulation of macrophage plasticity in polarization, inflammasome activation, cytokine secretion, metabolism, phagocytosis, and the number of macrophages. In addition, autophagy has been shown to connect macrophages and heart cells. It is attributed to specific substrate degradation or signalling pathway activation by autophagy-related proteins. Referring to the latest reports, applications targeting macrophage autophagy have been discussed in CVDs, such as atherosclerosis, myocardial infarction, heart failure, and myocarditis. This review describes a novel approach for future CVD therapies.

PGAM5 degrades PDCoV N protein and activates type I interferon to antagonize viral replication.

IMPORTANCE: As a member of the δ-coronavirus family, porcine deltacoronavirus (PDCoV) is a vital reason for diarrhea in piglets, which can contribute to high morbidity and mortality rates. Initially identified in Hong Kong in 2012, the virus has rapidly spread worldwide. During PDCoV infection, the virus employs evasion mechanisms to evade host surveillance, while the host mounts corresponding responses to impede viral replication. Our research has revealed that PDCoV infection down-regulates the expression of PGAM5 to promote virus replication. In contrast, PGAM5 degrades PDCoV N through autophagy by interacting with the cargo receptor P62 and the E3 ubiquitination ligase STUB1. Additionally, PGAM5 interacts with MyD88 and TRAF3 to activate the IFN signal pathway, resulting in the inhibition of viral replication.

N protein of PEDV plays chess game with host proteins by selective autophagy.

Macroautophagy/autophagy is a cellular degradation and recycling process that maintains the homeostasis of organisms. The protein degradation role of autophagy has been widely used to control viral infection at multiple levels. In the ongoing evolutionary arms race, viruses have developed various ways to hijack and subvert autophagy in favor of its replication. It is still unclear exactly how autophagy affects or inhibits viruses. In this study, we have found a novel host restriction factor, HNRNPA1, that could inhibit PEDV replication by degrading viral nucleocapsid (N) protein. The restriction factor activates the HNRNPA1-MARCHF8/MARCH8-CALCOCO2/NDP52-autophagosome pathway with the help of transcription factor EGR1 targeting the HNRNPA1 promoter. HNRNPA1 could also promote the expression of IFN to facilitate the host antiviral defense response for antagonizing PEDV infection through RIGI protein interaction. During viral replication, we found that PEDV can, in contrast, degrade the host antiviral proteins HNRNPA1 and others (FUBP3, HNRNPK, PTBP1, and TARDBP) through its N protein through the autophagy pathway. These results reveal the dual function of selective autophagy in PEDV N and host proteins, which could promote the ubiquitination of viral particles and host antiviral proteins and degradation both of the proteins to regulate the relationship between virus infection and host innate immunity.Abbreviations: 3-MA: 3-methyladenine; ATG: autophagy related; Baf A1: bafilomycin A1; CALCOCO2/NDP52: calcium binding and coiled-coil domain 2; ChIP: chromatin immunoprecipitation; Co-IP: co-immunoprecipitation; CQ: chloroquine; DAPI: 4',6-diamidino-2-phenylindole; GPI: glycosyl-phosphatidylinositol; hpi: hours post infection; MARCHF8/MARCH8: membrane-associated ring-CH-type finger 8; MOI: multiplicity of infection; N protein: nucleocapsid protein; PEDV: porcine epidemic diarrhea virus; siRNA: small interfering RNA; TCID50: 50% tissue culture infectious doses.

Efficacy and safety of 755-nm picosecond alexandrite laser with topical tranexamic acid versus laser monotherapy for melasma and facial rejuvenation: a multicenter, randomized, double-blinded, split-face study in Chinese patients.

To compare the efficacy and safety of 755-nm picosecond alexandrite laser and topical tranexamic acid (TTA) combination therapy with laser monotherapy, for the treatment of melasma and facial rejuvenation. This multicenter, randomized, double-blinded, split-face study enrolled 37 patients who presented with melasma and photoaging. Facial halves were randomized to receive either laser and TTA combination therapy or laser monotherapy. Three treatments were delivered at 4-5 weeks intervals. Patients were followed up for 1, 3, and 6 months post-final treatment and evaluated by blinded investigators for hemi-Melasma Area and Severity Index (hemi-MASI), facial dyschromia, skin texture, laxity, and rhytids. Daily diaries rating healing progress for 7 days posttreatment and satisfaction grading were performed by all patients. Adverse events were recorded. Thirty-six patients completed the follow-up. Compared with the baseline, hemi-MASI, dyschromia, and skin texture on both halves improved significantly through the follow-up (p = 0.000). A significant difference in hemi-MASI and dyschromia between combination therapy halves and monotherapy halves was noticed at 1- and 3-month follow-ups (p < 0.05). The laser monotherapy halves displayed significantly less redness and sensitivity during the 7-day posttreatment recovery period (p < 0.05). Patients' satisfaction ratings for the combination therapy halves were higher than the monotherapy halves at 1-month follow-up (p < 0.05). No severe adverse events were observed. The picosecond alexandrite laser and TTA combination therapy demonstrated synergistic efficacy for hemi-MASI and dyschromia improvements over laser monotherapy. The optimization of the picosecond laser and TTA combination regimen needs further investigation.

Expression pattern Nanog is associated with the development of gestational trophoblastic neoplasia in Chinese women from Fujian province: a case-control study.

BACKGROUND: Gestational trophoblastic disease (GTD) is a group of interrelated but distinct diseases and has a serious impact on the reproductive health of women. To analyze the expression of Nanog in GTD and to evaluate its potential to predict the development of gestational trophoblastic neoplasia (GTN). METHODS: The study included 41 normal first-trimester placentas matched by gestational age to 53 regressed-hydatidiform-moles (rHMs), 56 malignant-HMs (mHMs) and 17 choriocarcinomas (CCAs) and evaluated the Nanog expression by immunohistochemistry. The chi-square test, ANOVA, Fisher's exact test and logistic regression were performed to assess the Nanog expression and clinical prognostic factors in GTD. RESULTS: Compared to normal placenta levels, the Nanog expression was increased in GTD samples (p < .05). In HMs, Nanog expression was positively correlated with serum ß-hCG levels,uterine size and theca-lutein cysts (p < .05). Compared with the low-risk metastatic group (Federation of Gynecology and Obstetrics (FIGO) score ≤ 6), the high-risk metastatic group (FIGO score >7) had higher Nanog expression (p = .030). Moreover, logistic regression analysis showed that the positive expression of Nanog had the highest risk of developing into GTN (OR = 4.764, p < .001). CONCLUSIONS: Nanog is an independent predictor of clinical outcomes. It can also be a reliable predictor for GTN development from GTD.

Novel turn-on fluorescence sensor for detection and imaging of endogenous H2S induced by sodium nitroprusside.

Currently, fluorescence analysis method has a good application in the detection and imaging of biomarkers and has become an important analytical method. Although there are many fluorescent probes for detecting hydrogen sulfide(H2S), they are mostly based on fluorophores which already existed, such as 1,8-naphthalimide, coumarin, rhodamine and their derivatives. Here, a new type of fluorescent molecule (BOTD) was synthesized and applied to the detection of H2S. The probe BOTD could quickly and sensitively detect H2S and turn on fluorescence. Moreover, the probe BOTD was successfully applied to the detection of exogenous and endogenous H2S in living cells, and may be expected to become a research tool for studying H2S-induced drugs.

A double-indole structure fluorescent probe for monitoring sulfur dioxide derivatives with distinct ratiometric fluorescence signals in mammalian cells.

Based on the addition reaction of the sulfur dioxide derivative to the CC double bond, the probe HDI was designed and synthesized. The two-channel fluorescent probe HDI changed from orange to colorless and the fluorescence changed from red to blue when the bisulfite was detected. And the probe responds rapidly to bisulfite within 2 min, with high sensitivity and specificity. In addition, the probe can be used to detect the concentration of bisulfite with a low detection limit (80 nM). Cytological experiments have also demonstrated that probe HDI has low cytotoxicity and could be used for ratiometric detection of sulfur dioxide derivatives in living cells.

Deciphering the Antitoxin-Regulated Bacterial Stress Response via Single-Cell Analysis.

Bacterial toxin-antitoxin (TA) systems, which are diverse and widespread among prokaryotes, are responsible for tolerance to drugs and environmental stresses. However, the low abundance of toxin and antitoxin proteins renders their quantitative measurement in single bacteria challenging. Employing a laboratory-built nano-flow cytometer (nFCM) to monitor a tetracysteine (TC)-tagged TA system labeled with the biarsenical dye FlAsH, we here report the development of a sensitive method that enables the detection of basal-level expression of antitoxin. Using the Escherichia coli MqsR/MqsA as a model TA system, we reveal for the first time that under its native promoter and in the absence of environmental stress, there exist two populations of bacteria with high or low levels of antitoxin MqsA. Under environmental stress, such as bile acid stress, heat shock, and amino acid starvation, the two populations of bacteria responded differently in terms of MqsA degradation and production. Subsequently, resumed production of MqsA after amino acid stress was observed for the first time. Taking advantage of the multiparameter capability of nFCM, bacterial growth rate and MqsA production were analyzed simultaneously. We found that under environmental stress, the response of bacterial growth was consistent with MqsA production but with an approximate 60 min lag. Overall, the results of the present study indicate that stochastic elevation of MqsA level facilitates bacterial survival, and the two populations with distinct phenotypes empower bacteria to deal with fluctuating environments. This analytical method will help researchers gain deeper insight into the heterogeneity and fundamental role of TA systems.

Cervical cancer screening using the Cervista high-risk human papillomavirus test: opportunistic screening of a hospital-based population in Fujian province, China.

OBJECTIVES: The Cervista® high-risk human papillomavirus (HR-HPV) test was evaluated as a primary screening method for cervical cancer in women aged ≥21 years and was compared with different screening and triage combinations. MATERIALS AND METHODS: A nested case-control study within the Fujian provincial Cervical Lesion Screening Cohorts was used to evaluate the Cervista test as the primary cervical screening method in a hospital-based population. Strategy 1 primarily screened using a cytology screen with HR-HPV testing used for triage. Strategy 2 primarily screened using cytology and HR-HPV co-testing. Strategy 3 primarily screened using HR-HPV testing and triaged HPV-positive women based on cytology. Strategy 4 primarily screened using HR-HPV testing and referred A9 pool HPV-positive women to colposcopy directly, whereas non-A9 HPV-positive women were triaged using cytology. RESULTS: There were 10,183 women included in this study; 16.49% (1677/10,183) were HR-HPV-positive, 9.52% had abnormal cytology, and 9907 women were normal during followup. A total of 276 women were diagnosed with cervical intraepithelial neoplasia 2 or worse (CIN2+), 197 with CIN3 or worse (CIN3+), and 70 with cervical cancer. Moreover, 10.15% (20/197) women who were CIN3+ were identified as cytology-negative, while 8.63% (17/197) were HR-HPV negative (P>0.05). The cumulative risk rate for HPV-/cytology- was 0.836 (95% CI, 0.424-1.648) in CIN3+ cases. Strategy 4 yielded the highest sensitivity for CIN2+ or CIN3+ and the lowest positive predictive value for CIN2+ or CIN3+ among the four screening strategies. CONCLUSION: The Cervista HR-HPV test can provide a reliable and sensitive clinical reference for the cervical cancer primary screen.

Potential role of the HOXD8 transcription factor in cisplatin resistance and tumour metastasis in advanced epithelial ovarian cancer.

Few studies have examined the potential transcription factor (TF) simultaneously associated with cisplatin resistance and metastasis in ovarian cancer. To assess a related mechanism, a 345-channel protein/DNA array and transcriptional activity ELISA were performed to compare the TF activities in the cisplatin-sensitive SKOV3 and cisplatin-resistant SKOV3-DDP cells and in HO-8910 and the homologous highly metastatic HO-8910PM cells. In SKOV3-DDP vs. SKOV3 cells, 43 TFs were up-regulated, while 31 were down-regulated. In HO-8910PM vs. HO-8910 cells, 13 TFs were up-regulated, while 18 were down-regulated. In these two models, 4 TFs (HOXD8(1), HOXD8(2), RB, RFX1/2/3) were simultaneously up-regulated, and 9 TFs (SRE, FKHR, Angiotensinogen ANG-IRE, Pax2, CD28RC/NF-IL2B, HLF, CPE, CBFB and c-Ets-1) were down-regulated. HOXD8 mRNA and protein expression levels measured by reverse transcription polymerase chain reaction and ELISA, respectively, were significantly higher in SKOV3-DDP and HO-8910PM than in their corresponding cell lines (both p < 0.05). In 52 cases of different ovarian disease, the patients with recurrent and cisplatin-resistant ovarian cancer had higher expression levels of HOXD8 than patients with primary malignant tumours (p = 0.018, p = 0.001) or benign tumours (p = 0.001, p < 0.001). Taken together, these results suggest that HOXD8 is potentially associated with both cisplatin resistance and metastasis in advanced ovarian cancer.

Knowledge and Attitudes Regarding HPV and Vaccination Among Chinese Women Aged 20 to 35 Years in Fujian Province: A Cross-Sectional Study.

The use of the human papillomavirus (HPV) vaccine was recently approved in Mainland China. This study determined the knowledge and attitudes of young women aged 20 to 35 years in Fujian Province, China, with regard to HPV and vaccination and explored the potential factors influencing their attitudes toward HPV vaccination. This was a cross-sectional study that collected data regarding the knowledge on and attitudes toward HPV and vaccination using questionnaires. Furthermore, the prevalence of HPV was determined from the sampled participants. A total of 1001 young women were included in the survey. This study demonstrated that the HPV prevalence rate was 15.7% (157/1001). Among all patients, 44.9% (n = 449) had heard of HPV; however, detailed knowledge about HPV was lacking. The majority (83.7%) expressed a willingness to be vaccinated. Specifically, knowledge of the dangers of HPV infection was significantly associated with the willingness to be vaccinated. In this study, women cited some concerns and expressed high expectations for the HPV vaccine, but the costs of vaccination reduced their willingness to be vaccinated. This study found that most patients did not have a detailed knowledge of HPV. Thus, there is a need for continued HPV promotion and education efforts, especially on the dangers of HPV infection, among young women aged 20 to 35 years in Fujian Province, China. Furthermore, it is important to subsidize the costs of vaccination for promoting vaccination campaigns in China.

Regulation of matriptase and HAI-1 system, a novel therapeutic target in human endometrial cancer cells.

The effects of specific and non-specific regulation of matriptase on endometrial cancer cells in vitro were investigated. Messenger ribonucleic acid (mRNA) and protein expression of matriptase and hepatocyte growth factor activator inhibitor-1 (HAI-1) in RL-952, HEC-1A, and HEC-1B endometrial cancer cells were detected by real-time quantitative PCR (RT-qPCR) and western blot. The cells were infected with lentivirus-mediated small-interfering RNA (siRNA) targeted on matriptase (MA-siRNA) or treated with different cisplatin (DDP) concentrations. After treatment, invasion, migration, and cellular apoptosis were analyzed. Matriptase mRNA and protein expression significantly decreased to 80% after infection with MA-siRNA (P < 0.01), and scratch and trans-well chamber assays showed significant inhibition of invasiveness and metastasis. Upon incubation with cisplatin at concentrations higher than the therapeutic dose for 24 h, the expressions of matriptase and HAI-1 significantly decreased (P < 0.001). Moreover, the invasiveness, metastasis, and survival rate of HEC-1A and RL-952 endometrial cancer cells were significantly decreased (P < 0.001) due to the down-regulation of matriptase and HAI-1 upon increasing cisplatin concentration. However, a slight increase in matriptase and HAI-1 expression was observed in cells treated with low cisplatin concentration (P = 0.01). Moreover, matriptase expression was associated with metastasis and invasiveness. Down-regulation of matriptase by specific Ma-SiRNA or non-specific cisplatin in matriptase/HAI-1-positive endometrial cancer cells showed promising therapeutic features.

Clinical validation of the PCR-reverse dot blot human papillomavirus genotyping test in cervical lesions from Chinese women in the Fujian province: a hospital-based population study.

OBJECTIVE: To determine the clinical significance of the polymerase chain reaction (PCR)-reverse dot blot (RDB) human papillomavirus (HPV) genotyping assay in cervical cancer screening. METHODS: A total of 10,442 women attending the Fujian Provincial Maternity and Children's Health Hospital were evaluated using the liquid-based cytology (thinprep cytologic test [TCT]) and the PCR-RDB HPV test. Women with HPV infection and/or abnormal cytology were referred for colposcopy and biopsy. For HPV DNA sequencing, 120 specimens were randomly selected. Pathological diagnosis was used as the gold standard. RESULTS: Using the PCR-RDB HPV test, overall HPV prevalence was 20.57% (2,148/10,442) and that of high-risk (HR)-HPV infection was 18.68% (1,951/10,442). There was 99.2% concordance between HPV PCR-RDB testing and sequencing. In this studied population, the most common HR-HPV types were HPV-16, -52, -58, -18, -53, -33, and -51, rank from high to low. HPV-16, -18, -58, -59, and -33 were the top 5 prevalent genotypes in cervical cancer but HPV-16, -18, -59, -45, and -33 were the top 5 highest risk factors for cancer (odds ratio [OR]=34.964, 7.278, 6.728, 6.101, and 3.658; all p<0.05, respectively). Among 10,442 cases, 1,278 had abnormal cytology results, of which, the HR-HPV positivity rate was 83.02% (1,061/1,278). To screen for cervical cancer by PCR-RDB HPV testing, when using CIN2+, CIN3+, and cancer as observed endpoints, the sensitivity was 90.43%, 92.61%, and 94.78% and the negative predictive value (NPV) was 99.06%, 99.42%, and 99.78%, respectively. PCR-RDB HPV and TCT co-testing achieved the highest sensitivity and NPV. CONCLUSION: For cervical cancer screening, the PCR-RDB HPV test can provide a reliable and sensitive clinical reference.

Expression patterns of maspin and mutant p53 are associated with the development of gestational trophoblastic neoplasia.

Gestational trophoblastic disease (GTD) is a group of conditions that originate from the abnormal proliferation of trophoblastic cells. GTDs encompass hydatidiform moles (HMs) and gestational trophoblastic neoplasia (GTN). GTNs are a group of malignant diseases that require chemotherapy, or more aggressive treatment. There is a requirement for more tumor markers to predict the development of GTN from HMs. The current study evaluated the expression of maspin and tumor protein p53 (p53) in GTD, and their role in predicting the development of GTN. Expression of maspin and mutant p53 (m-p53) was detected by immunohistochemistry in 48 normal first trimester placentas, matched for gestational age to 49 HMs that regressed, 39 malignant HMs and 11 invasive moles or choriocarcinomas. Spearman's rank correlation analysis and logistic regression were performed on the expression patterns of maspin and m-p53, and on the clinical prognostic factors in GTD. Compared with normal placenta levels, the expression levels of maspin were decreased, whereas the expression levels of m-p53 were increased in GTDs (P<0.05). The expression levels of maspin and m-p53 in complete and partial HMs were not significantly different (P>0.05). In HMs, maspin expression was inversely correlated with serum ß human chorionic gonadotropin, uterine size and diameter of theca-lutein cysts; however, m-p53 expression demonstrated a positive correlation with these factors (all P<0.05). Compared with the high-risk metastatic group (FIGO score ≥7), the low-risk group (FIGO score <7) exhibited a higher rate of positive maspin expression (P=0.041), and the frequency of positive m-p53 expression was significantly higher in patients with an advanced FIGO stages (FIGO stage ≥III) compared with patients in early stages (FIGO stage ≤II; 87.9 vs. 58.8%; P=0.019). The combination of maspin negative expression with m-p53 positive expression had an 84% specificity value, 76% positive predictive value and 70% negative predictive value for the development of GTN. In conclusion, maspin-negative and m-p53-positive expression is associated with the development of GTN in HMs.

Decreasing the ratio of matriptase/HAI­1 by downregulation of matriptase as a potential adjuvant therapy in ovarian cancer.

Tumor invasion and metastasis are complex biological processes. Matriptase and its endogenous inhibitor, hepatocyte growth factor activator inhibitor­1 (HAI­1) are involved in invasion and metastasis. To evaluate the ratio of matriptase/HAI­1 and their potential therapeutic value in ovarian cancer, HO­8910 human ovarian cancer cells and the homologous high­metastatic HO­8910PM cells were used as in vitro cellular models ovarian cancer. The invasive and metastatic abilities, and the expression of matriptase and HAI­1 in these cells were detected using scratch assays, Transwell chamber assays, reverse transcription­quantitative polymerase chain reaction, western blotting and fluorescent immunocytochemistry. Following infection with lentivirus­mediated matriptase­targeting small interfering RNA (siRNA), cell cycle progression and apoptosis were also analyzed. The migration distance and number of invading HO­8910PM cells were significantly increased compared with HO­8910 cells. HO­8910PM cells exhibited a significantly higher ratio of matriptase/HAI­1 mRNA levels compared with HO­8910 cells (0.51 vs. 0.24, ~2.2 fold increase). Compared with HO­8910 cells, the matriptase mRNA level was increased by ~3.6 fold in HO­8910PM cells, whereas the HAI­1 mRNA level was increased by ~1.7 fold. Similar increases in protein expression levels were also observed in HO­8910PM cells compared with HO­8910 cells. Migration and invasiveness were positively correlated with matriptase expression level (r=0.994, P<0.01) and the ratio of matriptase/HAI­1 (r=0.929, P<0.01). Downregulation of matriptase using siRNA resulted in inhibition of the invasive and metastatic abilities of HO­8910PM cells, cell cycle arrest in the G0/G1 phase and increased apoptosis. The present study demonstrated that ovarian cancer cell metastasis and invasion were more dependent on upregulation of matriptase levels than downregulation of HAI­1. Matriptase may be a potential adjuvant therapeutic target for inhibiting ovarian cancer invasion and metastasis.

Specific detection of live Escherichia coli O157:H7 using tetracysteine-tagged PP01 bacteriophage.

Sensitive and rapid detection of Escherichia coli O157:H7, one of the most notorious bacterial pathogens, is urgently needed for public health protection. Yet, the existing methods are either lack of speed or limited in discriminating viable and dead cells. Using a recombinant bacteriophage, here we report the development of a rapid and sensitive method for live E. coli O157:H7 detection. First, the wild-type PP01 phage was engineered with a tetracysteine (TC)-tag fused with the small outer capsid (SOC) protein. Then, this PP01-TC phage was used to inoculate bacterial sample for 30min. Specific infection and rapid replication of PP01-TC phage in viable E. coli O157:H7 host cell yields a large number of progeny phages with capsids displaying TC tags that can be fluorescently labeled by a membrane permeable biarsenical dye (FlAsH). The bright green fluorescence of single E. coli O157:H7 cells can be readily detected by flow cytometry (FCM) and fluorescence microscopy. High specificity of the assay was verified with seven other bacterial strains. Practical application in E. coli O157:H7 detection in drinks was successfully demonstrated with artificially contaminated 100% apple juice. In less than three hours (including sample preconcentration) and with 40mL of sample volume, as low as 1cfu/mL E. coli O157:H7 can be detected in the presence of large excess of other nontarget bacteria via fluorescence microscopic measurement. The as-developed TC-PP01-FlAsH approach shows a great potential in the safeguard of liquid food products by providing rapid, sensitive, and specific detection of live E. coli O157:H7.

[Expression and significance of matriptase in ovarian cancer cells with diverse metastatic potential].

OBJECTIVE: To study the expression and significance of matriptase in different metastatic potential of human ovarian cancer cells. METHODS: High-metastatic human ovarian cancer cell HO8910PM and ovarian cancer cell HO8910 were collected.The ability of metastatic of the former was stronger than that of the latter. Compared the ability of invasion and migration in HO8910PM and HO8910 by scratch assay and by millicell chamber artificial reconstituted basement membrane invasion assay. Detected the matriptase mRNA and protein expression levels in HO8910PM and HO8910 through reverse transcription(RT)-PCR and immunocytochemistry methods. RESULTS: The 24 hours' migration distance (347 ± 8) µm of HO8910PM cells were significantly higher than that in HO8910 group (154 ± 10) µm (P < 0.01);The number of HO8910PM cells that penetrated the matrigel after 24 hours' incubation were significantly higher than that in HO8910 group (90.7 ± 2.1 vs 63.3 ± 1.5,P < 0.01). The expression of matriptase mRNA in HO8910PM cells was higher than that in HO8910 group (0.72 ± 0.03 vs 0.38 ± 0.04,P < 0.01). The migration was positively correlated with the matriptase mRNA expression levels (r = 0.992, P < 0.01); and the invasiveness was also positively correlated with the matriptase mRNA expression levels (r = 0.973, P < 0.01). As far protein level,the expression of matriptase protein in HO8910PM cells was higher than that in HO8910 group (15.6 ± 0.8 vs 7.6 ± 1.3,P < 0.01). The migration was positively correlated with matriptase protein expression levels (r = 0.971, P < 0.01); And the invasiveness was also positively correlated with the matriptase protein expression levels (r = 0.958, P < 0.01). CONCLUSIONS: The relationship between the expression levels of matriptase and the metastatic of ovarian cancer cells may be correlative. The function of matriptase in ovarian cancer cells metastatic mechanism still need to be confirmed.