Murine cytomegalovirus employs the mixed lineage kinases family to regulate the spiral ganglion neuron cell death and hearing loss.

Cytomegalovirus (CMV)-induced sensorineural hearing loss (SNHL) is a worldwide epidemic. Recent studies have shown that the degree of spiral ganglion neuron (SGN) loss is correlated with hearing loss after CMV infection. We aimed to better understand the pathological mechanisms of CMV-related SGN death and to search for intervention measures. We found that both apoptosis and pyroptosis are involved in CMV-induced SGN death, which may be caused by the simultaneous activation of the p53/JNK and NLRP3/caspase-1 signaling pathways, respectively. Moreover, considering that mixed lineage kinase family (MLK1/2/3) are host restriction factors against viral infection and upstream regulators of the p53/JNK and inflammatory (including NLRP3-caspase1) signaling pathways, we further demonstrated that the MLKs inhibitor URMC-099 exhibited a protective effect against CMV-induced SGN death and hearing loss. These results indicate that MLKs signaling may be a key regulator and promising novel target for preventing apoptosis and even pyroptosis during the CMV infection of SGN cells and for treating hearing loss.

LncRNA TCTN2 Promotes the Malignant Development of Hepatocellular Carcinoma via Regulating mIR-1285-3p/ARF6 Axis.

BACKGROUND: Hepatocellular carcinoma (HCC) is one of the most life-threatening malignant diseases. TCTN2 protein participates in tumorigenesis and development. However, whether lncRNA TCTN2 is associated with HCC pathogenesis remains unclear. METHODS: The expression of lncRNA, TCTN2, miR-1285-3p, and ARF6 in HCC tissues and cells was detected by a quantitative Real-Time PCR (qRT-PCR) assay. lncRNA TCTN2-specific shRNA was transfected into HCC cells, and a functional investigation was performed. The direct interactions between lncRNA TCTN2 and miR-1285-3p and ARF6 were verified by dualluciferase reporter gene assay. A rescue experiment was performed to confirm the role of miR- 1285-3p/ARF6 in association with lncRNA TCTN2. RESULTS: LncRNA TCTN2 exhibited a high expression in HCC tumor tissues and cell lines. Knockdown of lncRNA TCTN2 suppressed cell proliferation and induced cell cycle arrest and apoptosis through regulating Cyclin D1/p21 and Bax/Bcl-2 signals. Meanwhile, the knockdown of lncRNA TCTN2 inhibited HCC cell migration and invasion through upregulating MMP2/MMP9. Mechanistic investigation revealed that lncRNA TCTN2 upregulated the expression of ARF6 via sponging miR-1285-3p. Rescue experiments indicated that miR-1285-3p inhibitor reversed the antitumor effects of lncRNA TCTN2 and ARF6 knockdown inhibited the progression of HCC. CONCLUSION: Our results suggested that the knockdown of lncRNA TCTN2 inhibited HCC development by regulating the miR-1285-3p/ARF6 axis, implying that the lncRNA TCTN2 is upregulated in HCC and may serve as a diagnostic biomarker in HCC. Furthermore, it may demonstrate an important value for the clinical treatment of patients with HCC.

Co-activation of Caspase-1 and Caspase-8 in CMV-induced SGN death by inflammasome-associated pyroptosis and apoptosis.

Cytomegalovirus (CMV) infection causes newborn deafness, and the death of the spiral ganglion neurons (SGNs) is crucial in determining the degree of CMV-related hearing loss. Therefore, understanding the psychopathology of CMV-related SGN loss is important for identifying targets and exploring treatment strategies. In this study, we found that pyroptosis and apoptosis, two inflammasome-related programmed cell death pathways, are involved in CMV-induced SGN death and are mainly regulated by activated caspase-1 and caspase-8. Moreover, suppressing inflammasome assembly by blocking apoptosis-associated speck-like protein containing a CARD (ASC) interaction inhibited the activation of both caspase-1 and caspase-8, rescued SGN death, and improved hearing loss in CMV-infected newborn mice. Therefore, we propose that ASC inflammasome might be a promising target for treating CMV-related SGN death and newborn hearing loss by inhibiting caspase-1 and caspase-8 activated pyroptosis and apoptosis.

MicroRNA-641 Inhibits Endometrial Cancer Progression via Targeting AP1G1.

MicroRNA-641 (miR-641) was significantly decreased in various cancers, but its roles in endometrial cancer (EC) remain unclear. We explored the influences of miR-641 on the EC cells. In our study, the miR-641 expression was reduced in EC cells. Overexpression of miR-641 inhibited viability and proliferation of HEC-1A and HECCL-1 cells by CCK-8 and colony formation assays. Additionally, flow cytometry revealed that overexpression of miR-641 could remarkably promote apoptosis and arrest the cell cycle at the G1 phase of HEC-1A and HECCL-1 cells. Besides, forced expression of miR-641 suppressed the migration and invasion of HEC-1A and HECCL-1 cells as evidenced by wound healing and transwell assay. Moreover, AP1G1 was confirmed as a target gene of miR-641 by StarBase prediction and DLR assay and their expressions were negatively correlated. Overexpression of AP1G1 neutralized the roles of miR-641 mimic on the viability, proliferation, apoptosis, and migration of HEC-1A and HECCL-1 cells. Our findings illustrated that miR-641 was reduced in the EC cells and AP1G1 antagonized the miR-641 mimic-induced inhibition of the EC progression in vitro. Therefore, miR-641 may emerge as an effective molecule for EC treatment.

UPLC/Q-TOF-MS profiling of phenolics from Canarium pimela leaves and its vasorelaxant and antioxidant activities

ABSTRACT Canarium pimela K.D. Koenig, Burseraceae, have a long history of use in the Chinese traditional medicine treatment of various ailments including hypertension, and our research team has reported the anti-hypertensive activity and delineated the mechanism involved in the action. The following research aims to evaluate the vasorelaxant and antioxidant activities of ethanol extract from C. pimela leaves and to analyze its chemical composition by ultra-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UPLC/Q-TOF-MS) that may correlate with their pharmacological activities. The results showed that pre-incubation of aortic rings with the extract (0.3, 1, 3, 10, 30 and 100 mg/l) significantly inhibited the contractile response of the rings to norepinephrine-induced contraction (p < 0.01or p < 0.05). Crude ethanol extract and refined ethanol extract showed a highest inhibitory effect against 2,2dipheyl-2-picrylhydrazyl hydrate scavenging activity (IC50 of crude ethanol extract = 15.42 ± 0.14 µg/ml and IC50 of refined ethanol extract = 5.72 ± 0.31 µg/ml) and 2,2′-azinobis (3-ethyl-benzothiazoline-6-sulphonic acid ammonium salt) (ABTS (IC50 of crude ethanol extract = 3.24 ± 0.18 µg/ml and IC50 of refined ethanol extract = 1.88 ± 0.07 µg/ml) scavenging activity, which was considerably higher than that reported for butylated hydroxytoluene and lower of that measured for ascorbic acid. Moreover, its chemical composition was analyzed by UPLC/Q-TOF-MS. Sixteen compounds including nine flavonoids, four tannins, two phenolic acids and one dianthrone were identified for the first time as constituents of this species. And of this, six major phenolic components were simultaneous quantitative analysis by HPLC-UV, chlorogenic acid is the major compounds in C. pimela leaves. These results indicate that the phenolic-rich extract of C. pimela leaves is a promising natural pharmaceutical for combating hypertension and oxidative stress.

A screening analysis of the GJB2 c.176 del 16 mutation responsible for hereditary deafness in a Chinese family.

OBJECTIVE: To determine whether a new-born child from a family carrying a deafness gene needs cochlear implantation to avoid dysphonia by screening and sequencing a deafness-related gene. RESULTS: Both screening and sequencing results confirmed that the new born child had a normal GJB2 gene despite the fact that she has a brother suffering from hearing loss triggered by an allelic GJB2 c.176 del 16 mutation. We cloned the GJB2 genes derived from their respective blood genomic DNA into GFP fused plasmids and transfected those plasmids into the 293T cell line to test for gene function. While the mutated GJB2 gene (GJB2 c.176 del 16) of her deaf brother was found to be unable to form the gap junction structure between two adjacent cells, the baby girl's GJB2 gene ran into no such problems. CONCLUSION: The screening and sequencing as well as the GJB2 gene function tests invariably showed results consistent with the ABR tested hearing phenotype, which means that the child, with a normal wild type GJB2 gene, does not need early intervention to prevent her from developing hearing loss and dysphonia at a later stage in life.

Inflammasome activation in mouse inner ear in response to MCMV induced hearing loss.

OBJECTIVE: To identify presence of inflammasome activated in mouse cochlea with sensorineural hearing loss (SNHL) caused by cytomegalovirus (CMV) infection. METHOD: MCMV was injected into the right cerebral hemisphere in neonatal BALB/c mice at 2000 pfu virus titers. Auditory brainstem responses (ABRs) were tested to evaluate hearing at 21 days. Histopathological studies were conducted to confirm localizations of MCMV infected cells in the inner ear. Expression of inflammasome related factors was assessed by immunofluorescence, Quantitative real-time PCR and Western blotting. RESULTS: In the mouse model of CMV induced SNHL, inflammasome related kinase Caspase-1 and downstream inflammatory factor IL-1ß and IL-18 were found increased and activated after CMV infection in the cochlea. These factors could further up-regulate expression of IL-6 and TNF-α. These inflammatory factors are neurotoxicity and may contribute to hearing impairment. Furthermore, we also detected significantly increased AIM2 protein that accumulated in the SGN of cochleae with CMV infection. SIGNIFICANCE: We have shown that inflammasome as a novel inherent immunity mechanism may contribute to hearing impairment. CONCLUSION: Our data indicate that imflammasome assemble in mouse inner ear in response to CMV infection. We have revealed a novel pathology event in CMV induced SNHL involving activation of inflammasome in mouse cochlea. Additionally, we have shown that inflammasome may be a novel target for prevention and treatment of CMV related SNHL.

Novel-designed iliac branch stent graft for internal iliac artery reconstruction during aneurysm repair.

BACKGROUND: Iliac branch stent grafts (IBSGs) have been used for internal iliac artery preservation during aneurysm repair. However, current available branch iliac stent grafts used in Western countries are not suitable for many patients in Asia because of shorter common iliac arteries (CIA). The aim of this study was to evaluate the efficacy of a novel-designed IBSG in preservation of internal iliac artery during endovascular aneurysm repair in Chinese. METHODS: Eleven male patients (range, aged 65-80 years) underwent endovascular repair with 15 IBSGs, including 4 bilateral repairs between January 2011 and December 2012. The median abdominal aortic aneurysm diameter was 50 mm and the common iliac diameter was 38 mm. All patients received computed tomography angiography (CTA) before discharge and every 3 months afterward. RESULTS: The stent-graft deployment was technically successful in all cases. Seven of the 11 patients had uneventful procedures and the rest 4 complicated with fever, renal insufficiency, and groin hematoma. The median length of hospitalization was 7 days. Perioperative mortality was zero. Follow-up documented that 1 patient died at 14 months after surgery because of an unrelated cause. Two patients were found to have a type I and III endoleak, respectively, then successful treatments with internal iliac artery extension stent grafts were applied at 6 months. The overall primary patency was 86.7%. Follow-up CTA showed aneurysm shrinkage in all patients. CONCLUSIONS: These novel-designed IBSGs are safe and effective in preservation of internal iliac artery. It provides an alternative option for endovascular repair of the iliac aneurysms with short CIA, which is more common in Asian population.

Open and endovascular repair of primary mycotic aortic aneurysms: a 10-year single-center experience.

PURPOSE: To report early and midterm outcomes after open or endovascular repair of primary mycotic aortic aneurysms treated over a 10-year period in a single center. METHODS: The records of all 12 patients (10 men; 72.9 years, range 59-83) treated for primary mycotic aortic aneurysms from September 2001 to December 2010 were retrospectively reviewed. The aneurysms were located in the abdominal aorta in 10 cases and in the thoracic aorta in 2. Preoperative signs of infection, such as leukocytosis or elevated C-reactive protein, were found in all patients, and fever was apparent in 7. Three patients had primary open surgery with extensive debridement and extra-anatomical bypass, while 9 patients underwent endovascular aneurysm repair. At the time of operation, 10 mycotic aneurysms were already ruptured. All patients were prescribed lifelong antibiotics after discharge. RESULTS: Positive microbial cultures were found in 8 patients, including Salmonella species in 2, S. aureus in 3, E. coli in 1, and Streptococcus in 1. Mean follow-up was 29.9 months (range 1-98). Five patients took lifelong oral antibiotics after discharge with a mean medication duration of 17 months (range 1-65). Two of the 3 open surgery patients died (1 early). In the 9 endovascular repair patients, there was no early mortality, but 1 patient died at 6 months of an unknown cause. Additionally, 6 patients had a late relapse and underwent either secondary open surgical debridement (n=2) or computed tomography-guided drainage and antibiotic flush; 3 of the 6 died. CONCLUSION: Endovascular aneurysm repair is a reasonable short-term management for patients with hemodynamic instability or high surgical risk. However, the late relapse rate after endovascular repair was very high in this series, despite adjunctive drainage and aggressive antibiotic treatment.

Heat shock protein 90 mediates cytoprotection by H2S against chemical hypoxia-induced injury in PC12 cells.

1. Increasing evidence indicates that hydrogen sulphide (H2S) may serve as an important biological cytoprotective agent. Heat shock protein (Hsp) 90 can attenuate stress-induced injury. However, whether Hsp90 mediates the cytoprotective effect of H2S against chemical hypoxia-induced injury in PC12 cells is not known. 2. In the present study, CoCl2 (a chemical hypoxia mimetic) was used to treat PC12 cells to create a model of chemical hypoxia. To explore the role of Hsp90 in the cytoprotection afforded by H2S against chemical hypoxia-induced injury, 2 µmol/L 17-allylaminogeldanamycin (17-AAG), a selective inhibitor of Hsp90, was administered for 30 min prior to preconditioning with 400 µmol/L NaHS, followed by chemical hypoxia. 3. Cobalt chloride reduced cell viability (by 52.7 ± 1.5%), increased PC12 cell apoptosis (by 42.1 ± 1.5%), induced reactive oxygen species (ROS) by 3.79% compared with control and induced the dissipation of mitochondrial membrane potential (MMP) by 2.56% compared with control. 4. Pretreatment of PC12 cells with 100-400 µmol/L sodium hydrosulphide (NaHS), an H2S donor, for 3 h prior to exposure to 600 µmol/L CoCl2 provided significant, concentration-dependant protection to PC12 cells against CoCl2-induced cytotoxicity. Specifically, pretreatment of PC12 cells with 400 µmol/L NaHS decreased apoptosis to 16.77 ± 1.77% and blocked the CoCl2-induced increase in ROS production and loss of MMP. 5. At 400 µmol/L, NaHS upregulated Hsp90 in a time-dependant manner (over the period 0-180 min). In addition to its effects on Hsp90 expression, NaHS pretreatment of PC12 cells augmented the overexpression of Hsp90 induced by 600 µmol/L CoCl2 by 1.38-fold (P < 0.01). 6. Treatment of PC12 cells with 2 µmol/L 17-AAG for 30 min prior to NaHS pretreatment blocked the overexpression of Hsp90 induced by NaHS preconditioning, as evidenced by decreased cell viability (by 54.2 + 1.2%; P < 0.01), increased PC12 cell apoptosis (by 36.6 ± 1.2%; P < 0.01) and increasing ROS production. 7. The findings of the present study provide novel evidence that Hsp90 mediates H2S-induced neuroprotection against chemical hypoxia-induced injury via anti-oxidant and anti-apoptotic effects.

Rapid mycoplasma culture for the early diagnosis of Mycoplasma pneumoniae infection.

Early diagnosis of Mycoplasma pneumoniae (Mp) plays a pivotal role in its management. We evaluated the role of rapid culture method in early diagnosis of Mp infection and discussed the potential impact factors. A total of 2,600 patients with acute respiratory infection were included, and their pharyngeal swab samples were prepared for Mp rapid culture based on selective Mp fluid culture medium. The clinical contributing factors related to Mp infection were also explored. The positive rate of Mp culture in females was 41.75%, which was higher than that for males (37.63%). Mp infections were incidental to the children and elderly. The positive rates of Mp culture were higher in children aged 3-5 years and adults older than 70 years (54.05 and 31.48%, respectively), compared with other ages. In addition, Mp infection frequently occurred in winter (December-February) and spring (March-May), with significantly higher positive rates of Mp culture by 40.02 and 42.89 vs. 32.15 and 33.50% in summer (June-August) and autumn (September-November), respectively. The positive rate of rapid culture for Mp was slightly higher than that by Mp antibody assay, but the diagnostic accordance was well between these two methods (P>0.05). Furthermore, clinical common symptoms of respiratory tract infection, such as fever, cough, and expectoration, were not found specific in Mp infection, suggesting that they were not independent prognostic predictors for Mp infection. Therefore, rapid culture based on the Mp pathogen detection would have important clinical application for the early diagnosis of Mp infection.

[Massage manipulation of supplementing marrow and kneading tendon in treating 30 children with spastic cerebral palsy].

OBJECTIVE: To observe the therapeutic effect of massage with manipulation of supplementing marrow and kneading tendon (SMKT) on spastic cerebral palsy (CP). METHODS: A total of 60 children with CP were randomly assigned to the treatment group and the control group equally. All were treated with rehabilitation training, but massage with SMKT was carried out additionally for those in the treatment group, five times every week and 3 months as a therapeutic course. Clinical efficacy was assessed adopting the gross motor function measurement (GMFM-66) and the revised Ashworth scale (MAS) before and after treatment. RESULTS: All children showed significant improvements in GMFM-66 after treatment. Compared with baseline, the improvement was statistically significant (P <0.01). Significant difference was also found between the 2 groups in MAS and in GMFM scores after treatment (P <0.05). CONCLUSION: Massage with SMKT manipulation shows a better effect than rehabilitation training therapy alone in treating spastic CP.