Seasonal shift in gut microbiome diversity in wild Sichuan takin (Budorcas tibetanus) and environmental adaptation.

In this study, we investigated the change in microbiome composition of wild Sichuan takin (Budorcas tibetanus) during winter and spring and analyzed the physiological implications for such changes. Diversity analyses of the microbiome (average 15,091 high-quality reads per sample) in 24 fecal samples (15 from winter, 9 from spring) revealed that spring samples had higher species diversity and were compositionally different from winter samples (P < 0.05). Taxonomic composition analysis showed that the relative abundance increased in spring for Patescibacteria (2.7% vs. 0.9% in winter, P < 0.001) and Tenericutes (1.9% vs. 1% in winter, P < 0.05). Substantial increases in relative abundance of Ruminococcaceae and Micrococcaceae were identified in spring and winter, respectively. Mann-Whitney U and ANCOM identified seven differentially abundant genera: Enterococcus, Acetitomaculum, Blautia, Coprococcus 1, Lachnospiraceae UCG 008, Ruminococcus 2 and Ralstonia. All seven genera were significantly more abundant in spring (average 0.016-1.2%) than winter (average 0-0.16%), with the largest difference found in Ruminococcus (1.21% in spring vs. 0.16% in winter). The other six genera were undetectable in winter. Functional prediction and pathway analysis revealed that biosynthesis of cofactors (ko01240) had the highest gene count ratios in the winter, followed by the two-component system (ko02020). Seasonal variation affects the gut microbiomes in wild Sichuan takins, with winter associated with lower species diversity and spring with enrichment of cellulose-degrading genera and phytopathogens. Such changes were crucial in their adaptation to the environment, particularly the difference in food abundance.

Vessels that encapsulate tumor clusters (VETC) pattern predicts the efficacy of adjuvant TACE in hepatocellular carcinoma.

PURPOSE: Postoperative adjuvant trans-catheter arterial chemoembolization (TACE) is regarded as a common strategy for hepatocellular carcinoma (HCC) patients at a high risk of recurrence. However, there are currently no clinically available biomarkers to predict adjuvant TACE response. Vessels that encapsulate tumor clusters (VETC) can be used as an independent predictor of HCC prognosis. In this study, we aimed to explore whether the VETC pattern could predict adjuvant TACE benefit. METHODS: Vascular pattern and HIF-1α expression were detected in immunohistochemistry. The survival benefit of adjuvant TACE therapy for patients with or without VETC pattern (VETC+ /VETC-) was evaluated. RESULTS: The adjuvant TACE therapy obviously improved the TTR and OS in VETC+ patients, while adjuvant TACE therapy could not benefit from VETC- patients. Univariate and multivariate analysis revealed that adjuvant TACE therapy significantly improved the TTR and OS in VETC+ patients, but not in VETC- patients. In addition, the VETC+ , but not VETC- , patients could benefit from adjuvant TACE therapy in patients with high-risk factors of vascular invasion, larger tumor or multiple tumor. The mechanistic investigations revealed that the favorable efficacy of adjuvant TACE on VETC+ patients, but not VETC- ones, may be not due to the activation of HIF-1α pathway. CONCLUSION: The VETC pattern may represent a novel and reliable factor for selecting HCC patients who may benefit from adjuvant TACE therapy, and the combination of VETC pattern and tumor characteristics may help stratify patients' outcomes and responses to adjuvant TACE therapy.

Targeting USP1-dependent KDM4A protein stability as a potential prostate cancer therapy.

The histone demethylase lysine-specific demethylase 4A (KDM4A) is reported to be overexpressed and plays a vital in multiple cancers through controlling gene expression by epigenetic regulation of H3K9 or H3K36 methylation marks. However, the biological role and mechanism of KDM4A in prostate cancer (PC) remain unclear. Herein, we reported KDM4A expression was upregulation in phosphatase and tensin homolog knockout mouse prostate tissue. Depletion of KDM4A in PC cells inhibited their proliferation and survival in vivo and vitro. Further studies reveal that USP1 is a deubiquitinase that regulates KDM4A K48-linked deubiquitin and stability. Interestingly, we found c-Myc was a key downstream effector of the USP1-KDM4A/androgen receptor axis in driving PC cell proliferation. Notably, upregulation of KDM4A expression with high USP1 expression was observed in most prostate tumors and inhibition of USP1 promotes PC cells response to therapeutic agent enzalutamide. Our studies propose USP1 could be an anticancer therapeutic target in PC.

Metagenomic analysis of Sichuan takin fecal sample viromes reveals novel enterovirus and astrovirus.

The Sichuan takin inhabits the bamboo forests in the Eastern Himalayas and is considered as a national treasure of China with the highest legal protection and conservation status considered as vulnerable according to The IUCN Red List of Threatened Species. In this study, fecal samples of 71 Sichuan takins were pooled and deep sequenced. Among the 103,553 viral sequences, 21,961 were assigned to mammalian viruses. De novo assembly revealed genomes of an enterovirus and an astrovirus and contigs of circoviruses and genogroup I picobirnaviruses. Complete genome sequencing and phylogenetic analysis showed that Sichuan takin enterovirus is a novel serotype/genotype of the species Enterovirus G, with evidence of recombination. Sichuan takin astrovirus is a new subtype of bovine astrovirus, probably belonging to a new genogroup in the genus Mamastrovirus. Further studies will reveal whether these viruses can also be found in Mishmi takin and Shaanxi takin and their pathogenic potentials.

Quantitative analysis of gadoxetic acid-enhanced magnetic resonance imaging predicts histological grade of hepatocellular carcinoma.

OBJECTIVE: To confirm the histological grade of hepatocellular carcinoma (HCC) by gadoxetic acid-enhanced MRI. METHODS: Ninety-five HCC patients underwent gadoxetic acid-enhanced MRI before surgical intervention. The correlations among the signal absolute enhancement, contrast enhancement ratio (CER) and tumor histological grade were analyzed. RESULTS: The correlation between CER of tumor-to-liver and the grades of tumor differentiation is the most significant negative. The k-value for the CER of tumor-to-liver and histopathologic analysis is 0.62, which gives evidence of good agreement. CONCLUSION: The quantitative analysis of gadoxetic acid-enhanced MRI can predict the histological grades of small HCCs.

Role of New Nature Reserve in Assisting Endangered Species Conservation - Case Study of Giant Pandas in the Northern Qionglai Mountains, China.

The creation of nature reserves is the most direct way to save endangered species populations and their habitat. Development of the giant panda (Ailuropoda melanoleuca) nature reserve network in China was initiated in the 1960s, though the effort to create new reserves boomed considerably after the year 2000. Given this rapid development of protected areas in panda habitats, and the potential conflicting interests between conservation administrations and local economic development, it is essential to assess the role of new nature reserves in the overall giant panda conservation effort and reserve network. We utilized data from national giant panda surveys conducted in 2000 and 2012 to compare the size, spatial use, and distribution of panda populations, as well as the habitat suitability and connectivity in the Northern Qionglai Mountains between the two survey years. Our results show that although the total giant panda population in the study area did not change remarkably, local changes did occur. Most notably, the population in Wolong Nature Reserve declined by 27.3% (N = 39) and the population in Caopo Nature Reserve increased by 71.4% (N = 29) over the 12-year study period. We also found habitat suitability and availability decreased in both Wolong (12.4%) and Caopo (7.4%), but that the relative density of giant pandas declined (19.2%) and increased (84.6%) at each site, respectively. The distance between centers of high IUA were more distant in 2012 (14.1±1.9km) than that in 2000 (6.1±0.9km; t = -7.4, df = 5, p = 0.001), showing a scattered spatial pattern. Habitat availability decreased by 42% within the corridor between the two reserves, however panda occurrences in the corridor increased 24.6%. Compared to the total number of encounters, the proportion of the corridor increased 45.76%. Our results show the importance and success of the newly established Caopo to the conservation of giant pandas, and how crucial it is to identify and repair reserve corridors. Furthermore, we propose criteria for future nature reserve network management and investment, which is applicable for other endangered species conservation practices.

A Comparison between Three-Dimensional Visualization Guided Hepatectomy and Ultrasonography Guided Radiofrequency Ablation in the Treatment of Small Hepatocellular Carcinoma within the Milan Criteria.

Background. Treatment selection for small hepatocellular carcinoma (sHCC) is controversial. We aimed to compare the outcomes of medical imaging three-dimensional visualization system (MI-3DVS) guided surgical resection (SR) and ultrasonography guided radiofrequency ablation (RFA) for sHCC. Methods. In total, 194 patients who underwent SR or RFA in our hospital between January 2006 and May 2010 were retrospectively enrolled. Overall survival (OS), recurrence-free survival (RFS), and postoperative complications were compared. Cox regression was used to estimate the benefits of MI-3DVS-guided SR on OS and RFS. Results. Ninety-two patients underwent SR and 102 underwent RFA. The SR group experienced more complications (41.3% versus 19.6%) and longer hospital stay (18.04 ± 7.11 versus 13.06 ± 5.59) (both p < 0.05). The 1-, 2-, 3-, 4-, and 5-year OS was 96.7%, 95.7%, 93.5%, 83.5%, and 61.1% in the SR group and 95.0%, 88.1%, 72.7%, 56.9%, and 39.5% in the RFA group. Corresponding RFS was 95.7%, 94.6%, 84.7%, 59.8%, and 40.2% in SR group and 91.2%, 80.3%, 60.5%, 32.3%, and 22.3% in RFA group. The 5-year OS and RFS were higher in SR group (both p < 0.001). Interestingly, there was no significance in OS and RFS among subgroups aged >60 years. Independent predictors of OS and RFS, respectively, were intervention (HR, 2.769 and 1.933), tumor number (HR, 5.128 and 3.903), and serum alpha-fetoprotein (AFP) (HR, 1.871 and 1.474) (all p < 0.05). Conclusions. MI-3DVS based hepatectomy should be considered primary treatment while RFA can be treated as alternative therapy for older patients. Intervention, tumor number, and AFP are independent predictors for both survival and recurrence.

GP73 is down-regulated in gastric cancer and associated with tumor differentiation.

BACKGROUND: Golgi protein 73 (GP73) is a type II Golgi transmembrane protein. It is over-expressed in several cancers, including hepatocellular carcinomas, bile duct carcinomas, lung cancer and prostate cancer. However, there are few reports of GP73 in gastric cancer. This study is aimed at investigating the expression of GP73 and its relationship with clinical pathological characters in gastric cancer. METHODS: GP73 mRNA level was determined by quantitative real-time RT-PCR in 41 pairs of matched gastric tumorous tissues and adjacent non-tumorous mucosal tissues. Western blotting was also performed to detect the GP73 protein level. GP73 protein expression was analyzed by immunohistochemistry in 52 clinically characterized gastric cancer patients and 10 non-tumorous gastric mucosal tissue controls. RESULTS: The mRNA and protein level of GP73 were significantly down-regulated in gastric tumorous tissues compared with the non-tumorous mucosal tissues. In non-tumorous mucosa, strong diffuse cytoplasmic staining can be seen in cells located at the surface of the glandular and foveolar compartment; while in tumorous tissues, the staining was much weaker or even absent, and mainly in a semi-granular dot-like staining pattern. The expression level of GP73 protein was associated with patients' gender and tumor differentiation. CONCLUSIONS: GP73 was normally expressed in non-tumorous gastric mucosa and down-regulated in gastric cancer. Its expression in gastric cancer was correlated with tumor differentiation.

[Expression of aldo-keto reductase family 1 member B10 in gastric cancer tissues and its clinical significance].

OBJECTIVE: To investigate the association of AKR1B10 expression in gastric cancer tissues with clinicopathologic features and prognosis of gastric cancer patients. METHODS: Real-time polymerase chain reaction (RT-PCR) was conducted to detect AKR1B10 mRNA expression in gastric cancer and adjacent gastric mucosa tissues (n=36). AKR1B10 protein expression was measured by immunohistochemistry in primary gastric cancer tissues (n=100) and non-tumorous gastric mucosa tissues (n=70). RESULTS: RT-PCR results confirmed that AKR1B10 was significantly down-regulated in gastric cancer tissues compared with that in paired adjacent mucosa [8.3% (3/36) vs. 91.7% (33/36), P=0.000]. Immunohistochemistry revealed that the percentage of AKR1B10 positive specimens in gastric carcinoma was lower than that in normal specimens [33.0% (33/100) vs. 92.9% (65/70), P=0.000]. The frequencies of positive AKR1B10 in patients was significantly correlated with tumor size (P=0.000), invasive depth (P=0.004), lymph node metastasis (P=0.028), distant metastasis (P=0.031) and TNM stages (P=0.000). The 5-year survival rate of positive AKR1B10 group was significantly higher as compared to negative group (60.6% vs. 32.8%, P<0.01). CONCLUSION: The down-regulation of AKR1B10 expression in gastric cancer may be associated with the progress of gastric cancer is suggestive of poor prognosis.

The MDM2 309T>G polymorphism and ovarian cancer risk: a meta-analysis of 1534 cases and 2211 controls.

BACKGROUND: Recently, there have been a number of studies on the association between MDM2 (Murine Double Minute 2) 309 polymorphism and ovarian cancer risk. However, the results of previous reports remain controversial and ambiguous. Thus, we performed a meta-analysis to explore more precisely the association between MDM2 309 polymorphism and the risk of ovarian cancer. METHODS: A meta-analysis was performed to examine the association between MDM2 309T>G polymorphism and ovarian cancer risk. Odds ratio (OR) and its 95% confidence interval (CI) were used for statistical analysis. RESULTS: Our publication search identified a total of 6 studies with 1534 cases and 2211 controls. No significant association was found between MDM2 309T>G polymorphism and ovarian cancer risk in total population analysis. In the subgroup meta-analysis by ethnicity, a negative association was shown in Asian subgroup (G vs. T OR = 0.774, 95% CI = 0.628-0.955, P = 0.017, P(het) = 0.327; GG vs. TT: OR = 0.601, 95% CI = 0.395-0.914, P = 0.017, P(het) = 0.417; dominant model TG+GG vs. TT: OR = 0.661, 95% CI = 0.468-0.934, P = 0.019, P(het) = 0.880), and no significant association in any genetic models among Caucasians was observed. CONCLUSIONS: This meta-analysis provides evidence for the association between MDM2 309 polymorphism and ovarian cancer risk, supporting the hypothesis that MDM2 SNP309 G allele acts as an important ovarian cancer protective factor in Asians but not in Caucasians.

REG4 contributes to the invasiveness of pancreatic cancer by upregulating MMP-7 and MMP-9.

Recent studies have shown that overexpression of regenerating gene family member 4 (REG4) is associated with the initiation and progression of pancreatic cancer. In our study, we explored the role of REG4 in the invasion of pancreatic cancer. Real-time PCR and Western blot analysis were used to determine REG4 expression in pancreatic cancer cell lines. An MTT assay was carried out to test the effect of REG4 on the growth of pancreatic cancer cells. The involvement of REG4 in cancer cell invasion was examined by Transwell invasion assay. Two MMPs, MMP-7 and MMP-9, were identified from a pool of candidate genes as being related to REG4-induced cell invasion by PCR and Western blotting. Immunohistochemistry was used to confirm the correlation between REG4 and the two MMPs. High expression of REG4 was found in BXPC-3 cells and its culture media. But in PANC-1 and ASPC-1 cell lines, REG4 expression levels were very low, and no detectable protein was found in the culture medium. The MTT and Transwell invasion assays showed that recombinant REG4 protein and BXPC-3 conditioned media significantly promoted the proliferation and invasiveness of pancreatic cancer cells. It was also shown that MMP-7 and MMP-9 are upregulated by REG4 induction using real-time PCR and Western blotting analysis. Immunohistochemical study further verified this result. In conclusion, REG4 promotes not only growth but also in vitro invasiveness of pancreatic cancer cells by upregulating MMP-7 and MMP-9.

Does a temperate ungulate that breeds in summer exhibit rut-induced hypophagia? Analysis of time budgets of male takin (Budorcas taxicolor) in Sichuan, China.

Mammals maximize fitness by optimizing time and energy allocation between reproduction and survival. Describing time budgets is a way to understand a species' constraints in energy allocation. We describe a time budget for male takin (Budorcas taxicolor) in Tangjiahe Nature Reserve, China, to better understand rut-induced hypophagia, which is frequently observed in temperate ungulates that breed in autumn or in winter. Observations generally occurred at two elevations (1200-1600m and 2600-3200m), using 20-min focal animal scan sampling from 2007 to 2009. Feeding behaviors accounted for the majority in takin's time budget (61.1%) during daylight hours, relative to the other observed behaviors, such as rest (14.1%), alert behavior (10.2%) and locomotion (6.8%). We found a negative correlation between feeding behavior and rutting behavior during the rutting season. A ratio of feeding time to resting time increased from pre-rut to rut, while resting behavior did not change significantly across seasons. These results suggest the "energy saving" hypothesis could explain reduced foraging in male takin during the rut, but aspects of the species biology suggest that hypotheses for rut-induced hypophagia developed for other temperate ungulates do not apply to takin. We suggest that the unusual summer rutting season of takin releases males from the energy constraints encountered by temperate ungulates that breed in the autumn and has other benefits for offspring survival. Further research should be conducted on ungulates that exhibit rut during the summer and tropical ungulates that might not experience limited food availability following the mating season to improve our understanding on rut-induced hypophagia.

Oridonin induces apoptosis in gastric cancer through Apaf-1, cytochrome c and caspase-3 signaling pathway.

AIM: To investigate the effect and mechanism of oridonin on the gastric cancer cell line HGC-27 in vitro. METHODS: The inhibitory effect of oridonin on HGC-27 cells was detected using the 3-(4, 5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay. After treatment with 10 µg/mL oridonin for 24 h and 48 h, the cells were stained with acridine orange/ethidium bromide. The morphologic changes were observed under an inverted fluorescence microscope. DNA fragmentation (a hallmark of apoptosis) and lactate dehydrogenase activity were examined using DNA ladder assay and lactate dehydrogenase-release assay. After treated with oridonin (0, 1.25, 2.5, 5 and 10 µg/mL), HGC-27 cells were collected for anexin V-phycoerythrin and 7-amino-actinomycin D double staining and tested by flow cytometric analysis, and oridonin- induced apoptosis in HGC-27 cells was detected. After treatment with oridonin for 24 h, the effects of oridonin on expression of Apaf-1, Bcl-2, Bax, caspase-3 and cytochrome c were also analyzed using reverse-transcript polymerase chain reaction (RT-PCR) and Western blotting. RESULTS: Oridonin significantly inhibited the proliferation of HGC-27 cells in a dose- and time-dependent manner. The inhibition rates of HGC-27 treated with four different concentrations of oridonin for 24 h (1.25, 2.5, 5 and 10 µg/mL) were 1.78% ± 0.36%, 4.96% ± 1.59%, 10.35% ± 2.76% and 41.6% ± 4.29%, respectively, which showed a significant difference (P < 0.05). The inhibition rates of HGC-27 treated with oridonin at the four concentrations for 48 h were 14.77% ± 4.21%, 21.57% ± 3.75%, 30.31% ± 4.91% and 61.19% ± 5.81%, with a significant difference (P < 0.05). The inhibition rates of HGC-27 treated with oridonin for 72 h at the four concentrations were 25.77% ± 4.85%, 31.86% ± 3.86%, 48.30% ± 4.16% and 81.80% ± 6.72%, with a significant difference (P < 0.05). Cells treated with oridonin showed typical apoptotic features with acridine orange/ethidium bromide staining. After treatment with oridonin, the cells became round, shrank, and developed small buds around the nuclear membrane while forming apoptotic bodies. Lactate dehydrogenase (LDH) release assay showed that after treated with 1.25 µg/mL and 20 µg/mL oridonin for 24 h, LDH release of HGC-27 caused by apoptosis increased from 22.94% ± 3.8% to 52.68% ± 2.4% (P < 0.001). However, the change in the release of LDH caused by necrosis was insignificant, suggesting that the major cause of oridonin-induced HGC-27 cell death was apoptosis. Flow cytometric analysis also revealed that oridonin induced significant apoptosis compared with the controls (P < 0.05). And the apoptosis rates of HGC-27 induced by the four different concentrations of oridonin were 5.3% ± 1.02%, 12.8% ± 2.53%, 28.5% ± 4.23% and 49.6% ± 3.76%, which were in a dose-dependent manner (P < 0.05). After treatment for 24 h, DNA ladder showed that oridonin induced a significant increase in DNA fragmentation in a dose-dependent manner. RT-PCR revealed that mRNA expression levels were up-regulated compared with the controls in caspase-3 (0.917 ± 0.103 vs 0.357 ± 0.019, P < 0.05), cytochrome c (1.429 ± 0.111 vs 1.002 ± 0.014, P < 0.05), Apaf-1 (0.688 ± 0.101 vs 0.242 ± 0.037, P < 0.05) and Bax (0.856 ± 0.101 vs 0.278 ± 0.027, P < 0.05) (P < 0.05), whereas down-regulated in Bcl-2 (0.085 ± 0.012 vs 0.175 ± 0.030, P < 0.05). Western blotting analysis also confirmed this result. CONCLUSION: Apoptosis of HGC-27 induced by oridonin may be associated with differential expression of Apaf-1, caspase-3 and cytochrome c, which are highly dependent upon the mitochondrial pathway.