Background: Colorectal cancer remains to be the third leading cause of cancer mortality rates. Despite the diverse effects of the miRNA cluster located in PVT1 of 8q24.21 across various tumors, the specific biological function in colorectal cancer has not been clarified. Methods: The amplification of the miR-1204 cluster was analyzed with the cBioPortal database, while the expression and survival analysis of the miRNAs in the cluster were obtained from several GEO databases of colorectal cancer. To investigate the functional role of miR-1204 in colorectal cancer, overexpression and silencing experiments were performed by miR-1204 mimic and inhibitor transfection in colorectal cancer cell lines, respectively. Then, the effects of miR-1204 on cell proliferation were assessed through CCK-8, colony formation, and Edu assay. In addition, cell migration was evaluated using wound healing and Transwell assay. Moreover, candidate genes identified through RNA sequencing and predicted databases were identified and validated using PCR and western blot. A Dual-luciferase reporter experiment was conducted to identify MASPIN as the target gene of miR-1204. Results: In colorectal cancer, the miR-1204 cluster exhibited high amplification, and the expression levels of several cluster miRNAs were also significantly increased. Furthermore, miR-1204 was found to be significantly associated with disease-specific survival according to the analysis of GSE17536. Functional experiments demonstrated that transfection of miR-1204 mimic or inhibitor could enhance or decrease cancer cell proliferation and migration. MASPIN was identified as a target of miR-1204. Additionally, the overexpression of MASPIN partially rescued the effect of miR-1204 mimics on tumorigenic abilities in LOVO cells. Conclusion: miR-1204 positioning in 8q24.21 promotes the proliferation and migration of colorectal cancer cells by targeting MASPIN.

The inflammatory responses involving infiltration and activation of liver macrophages play a vital role in acute liver failure (ALF). In the liver of ALF mice, cannabinoid receptor 2 (CB2R) is significantly upregulated on macrophages, while CB2R agonist GW405833 (GW) could protect against cell death in acute liver damage. In this study, we investigated the molecular mechanisms underlying the protective effects of GW against ALF in vivo and in vitro from a perspective of macrophage glycometabolism. Mice were pretreated with GW (10 mg/kg, i.p.), then were injected with D-GalN (750 mg/kg, i.p.) and LPS (10 mg/kg, i.p.) to induce ALF. We verified the protective effects of GW pretreatment in ALF mice. Furthermore, GW pretreatment significantly reduced liver macrophage infiltration and M1 polarization, and inhibited the release of inflammatory factors TNF-α and IL-1β in ALF mice. These protective effects were eliminated by CB2R antagonist SR144528 or in CB2R^−/− ALF mice. We used LPS-stimulated RAW264.7 cells as an in vitro M1 macrophage-centered model of inflammatory response, and demonstrated that pretreatment with GW (10 μM) significantly reduced glucose metabolism by inhibiting glycolysis, which inhibited LPS-induced macrophage proliferation and inflammatory cytokines release. We verified these results in a stable CB2R^−/− RAW264.7 cell line. Moreover, we found that GW significantly inhibited the expression of hypoxia inducible factor 1α (HIF-1α). Using a stable HIF-1α^−/− RAW264.7 cell line, we confirmed that GW reduced the release of inflammatory cytokines from macrophages and inhibited glycolysis by downregulating HIF-1α expression. In conclusion, activation of CB2Rs inhibits the proliferation of hepatic macrophages and release of inflammatory factors in ALF mice through downregulating HIF-1α to inhibit glycolysis.

Heilongjiang Province located in northeast China is a multi-ethnic region with people who have lived in cold conditions for several generations. Fatty acids are important to people with cold resistance. CPT1A encodes a protein that imports long-chain fatty acids into the mitochondria for fatty-acid oxidation. FADS is an essential enzyme for the synthesis of long-chain polyunsaturated fatty acids. In the present study, we investigated the distributions of three cold resistance-related SNPs (rs80356779 G > A in CPT1A, rs7115739 T > G in FADS3 and rs174570 C > T in FADS2) from six populations that included 1093 individuals who have lived in Heilongjiang Province for at least three generations. The frequencies of rs174570 and rs7115739 were different in our six north minorities compared to the Chinese Dai in Xishuangbanna (CDX) in southern China. All the SNPs in Hezhen were significantly different from other five studied populations. In addition, the genetic distribution of rs174570 in Daur was significantly different from Manchu and Korea, and the frequency of rs7115739 in Ewenki was significantly different from the other populations. The results also showed that the frequencies of the three SNPs in the six minorities were different from those of Greenlandic Inuit and Siberian population. Our results showed the distributions of the three cold resistance-related SNPs from six populations that included 1093 individuals in northern China. Distributions of the allele frequencies for the cold resistance-related SNPs in northern China were statistically different from those in southern China. These data help to establish the DNA genome database for the six populations and fully preserve existing minority genetic information.

Progressive familial intrahepatic cholestasis type 3 (PFIC3) is a hepatic disorder occurring predominantly in childhood and is difficult to diagnose. PFIC3, being a rare autosomal recessive disease, is caused by genetic mutations in both alleles ofABCB4, resulting in the disruption of the bile secretory pathway. The identification of pathogenic effects resulting from different mutations in ABCB4 is the key to revealing the internal cause of disease. These mutations cause truncation, instability, misfolding, and impaired trafficking of the MDR3 protein. Here, we reported a girl, with a history of intrahepatic cholestasis and progressive liver cirrhosis, with an elevated gamma-glutamyltransferase level. Genetic screeningviawhole exome sequencing found a novel homozygous missense mutationABCB4:c.1195G>C:p.V399L, and the patient was diagnosed with PFIC3. Various computational tools predicted the variant to be deleterious and evolutionary conserved. For functional characterization studies, plasmids, encodingABCB4wild-type and selected established mutant constructs, were expressed in human embryonic kidney (HEK-293T) and hepatocellular carcinoma (HepG2) cells.In vitroexpression analysis observed a reduced expression of mutant protein compared to wild-type protein. We found thatABCB4wild type was localized at the apical canalicular membrane, while mutant p.V399L showed intracellular retention. Intracellular mistrafficking proteins usually undergo proteasomal or lysosomal degradation. We found that after treatment with proteasomal inhibitor MG132 and lysosomal inhibitor bafilomycin A1, MDR3 expression of V399L was significantly increased. A decrease in MDR3 expression of mutant V399L protein may be a result of proteasomal or lysosomal degradation. Pharmacological modulator cyclosporin A and intracellular low temperature (30°C) treatment significantly rescued both the folding defect and the active maturation of the mutant protein. Our study identified a novel pathogenic mutation which expanded the mutational spectrum of theABCB4gene and may contribute to understanding the molecular basis of PFIC3. Therefore, genetic screening plays a conclusive role in the diagnosis of rare heterogenic disorders like PFIC3.

Lung ageing alters the intrinsic structure of the lung and pulmonary surfactant system and increases the mortality and morbidity due to respiratory diseases in elderly individuals. We hypothesized that lung ageing results from an insufficiency of type II alveolar epithelial cells (AECIIs) in the lung tissue. Sirtuin 3 (SIRT3) is a member of the sirtuin family of proteins that promote longevity in many organisms. Increased SIRT3 expression has been linked to an extended lifespan in humans. Hence, we speculated that the overexpression of SIRT3 may help to ameliorate lung senescence and improve AECII function. AECIIs were isolated from young and old patients with pneumothorax caused by pulmonary bullae. The expression of SIRT3, manganese superoxide dismutase and catalase, as well as cell function and senescence indicators of young and old AECIIs, were measured before and after SIRT3 overexpression. After SIRT3 overexpression, the aged state of old AECIIs improved, and anti-apoptotic activity, proliferation and secretion were dramatically enhanced. Surfactant protein C (SPC), which is secreted by AECIIs, reduces alveolar surface tension, repairs the alveolar structure and regulates inflammation. SPC deficiency in patients is associated with increased inflammation and delayed repair. SIRT3 deacetylated forkhead box O3a, thereby protecting mitochondria from oxidative stress and improving cell function and the senescent state of old AECIIs. These findings provide a possible direction for ageing-delaying therapies and interventions for diseases of the respiratory system.

Lynch syndrome (LS) is the most common hereditary colorectal cancer (CRCs) inherited in an autosomal-dominant manner. Here, we reported a multigeneration Chinese family clinically diagnosed with LS according to the Amsterdam II criteria. To identify the underlying causative gene for LS in this family, whole-exome sequencing (WES) was performed. A germline missense variant (c.2054C>T:p.S685F) in exon 18 of MLH1 was successfully identified by WES. Sanger sequencing verified the results of WES and also confirmed the cosegregation of the MLH1 missense variant in all affected members of the family including two unaffected family members. Bioinformatic tools predicted the identified MLH1 variant as deleterious. Immunohistochemistry (IHC) staining showed loss of MLH1 and PMS2 protein expression. In vitro expression analysis also revealed that the identified MLH1 missense variant (c.2054C>T:p.S685F) results in reduced expression of both MLH1 and PMS2 proteins. Based on the American College of Medical Genetics and Genomics (ACMG) guidelines, the missense mutation c.2054C>T in MLH1 was classified as a “pathogenic” variant. Two unaffected family members were later recommended for colonoscopy and other important cancer diagnostic inspections every 1-2 years as both were at higher risk of LS. In conclusion, our findings widen the genotypic spectrum of MLH1 mutations responsible for LS. This study increases the phenotypic spectrum of LS which will certainly help the clinicians in diagnosing LS in multigeneration families. This study also puts emphasis on the importance of genetic counselling for the benefit of asymptomatic carriers of MMR gene variants who are at higher risk of LS.