Development of anti-bacterial adhesion and antibacterial sulfobetaines modified chitosan/polyvinyl alcohol composite films as packaging materials.

Chitosan exhibits a wide source, non-toxic and biodegradable, and is the optimal functional raw material for preparing food packaging materials. However, the pure chitosan film has some disadvantages such as limited antibacterial activity and weak mechanical properties. In this study, sulfobetaines modified chitosan (CS-SBMA) was synthesized by grafting copolymerized betaine methacrylate sulfonate onto the chain of chitosan to improve the anti-bacterial adhesion and antibacterial properties of chitosan, aiming to develop antibacterial and anti-bacterial adhesion films based on CS-SBMA and polyvinyl alcohol (PVA) by the casting method. The structure of CS-SBMA was characterized by 1H NMR and FTIR. The appropriate proportion of CS-SBMA/PVA was determined to be 1/1 and 1/2, by characterizing the composite films with FTIR, XRD, SEM, mechanical, optical, and water resistance behaviors. In addition, CS-SBMA/PVA films showed excellent antibacterial, anti-bacterial adhesion and biofilm control function. The colonies number of E. coli and S. aureus on the surface of CS-SBMA/PVA 1/1 film decreased 94.15 % and 94.27 %, respectively, and 92.93 % of S. aureus and 94.87 % of E. coli colonies were inactivated within 60 min contact. These results indicate that CS-SBMA/PVA film exhibits potential antibacterial and anti-bacterial adhesion properties, which is suitable for food packaging materials.

Total flavonoids of Rhizoma drynariae improves tendon-bone healing for anterior cruciate ligament reconstruction in mice and promotes the osteogenic differentiation of bone mesenchymal stem cells by the ERR1/2-Gga1-TGF-ß/MAPK pathway.

BACKGROUND: Total flavonoids of Rhizoma drynariae (TFRD) is broadly used in the treatment of orthopedic diseases. Nevertheless, the effects and underlying mechanism of TFRD on tendon-bone healing after anterior cruciate ligament reconstruction (ACLR) remain unclear. METHODS: The ACLR mouse model was established. Hematoxylin and Eosin (HE) staining was used for histological analysis of tendon-bone healing. Western blot was utilized to detect the levels of osteogenic related factors (ALP, OCN, RUNX2). The viability and alkaline phosphatase (ALP) activity of bone mesenchymal stem cells (BMSCs) were determined by Cell Counting Kit-8 (CCK-8) and ALP assays. The interaction of estrogen related receptor alpha (ESRRA), estrogen related receptor beta (ESRRB), and golgi-localized γ-ear containing ADP ribosylation factor-binding protein 1 (Gga1) was detected by luciferase reporter assays. The levels of important proteins on the TGF-ß/MAPK pathway were measured by western blot. RESULTS: TFRD improved tendon-bone healing, restored biomechanics of ACLR mice and activated the TGF-ß/MAPK pathway. TFRD treatment also enhanced the viability and osteogenic differentiation of BMSCs in vitro. Then, we demonstrated that TFRD targeted ESRRA and ESRRB to transcriptionally activate Gga1 expression. Knockdown of ESRRA, ESRRB, or Gga1 suppressed the viability and osteogenic differentiation of TFRD-induced BMSCs, which was revealed to be restored by Gga1 overexpression. The overexpression of ESRRA, ESRRB, or Gga1 was demonstrated to promote the BMSC viability and osteogenic differentiation. TGF-ß1 treatment can reverse the impact of Gga1 inhibition on osteogenic differentiation in TFRD-induced BMSCs. CONCLUSION: TFRD improves tendon-bone healing in ACLR mouse models and facilitates the osteogenic differentiation of BMSCs through the ERR1/2-Gga1-TGF-ß/MAPK pathway, which might deepen our understanding of the underlying mechanism of TFRD in tendon-bone healing.

1064nm Nd:YAG laser promotes chondrocytes regeneration and cartilage reshaping by upregulating local estrogen levels.

Cartilage is frequently used as a scaffolds for repairing and reconstructing body surface organs. However, after successful plastic surgery, transplanted cartilage scaffolds often exhibit deformation and absorption over time. To enhance the shaping stability of cartilage scaffolds and improve patients' satisfaction after reconstructions, we employed the ear folding models in New Zealand rabbits to confirm whether the 1064nm neodymium-doped yttrium aluminum garnet (Nd:YAG) laser could promote cartilage reshaping. There was an increase in collagen and aromatase (Cyp19) expression within the ear cartilage after laser treatment. Moreover, we have found that the Cyp19 inhibitor can inhibit the laser's effect on cartilage shaping and reduce collagen and Cyp19 expression. The overall findings suggest that treatment with 1064nm Nd:YAG laser irradiation can enhance estrogen levels in local cartilage tissues by upregulating Cyp19 expression in chondrocytes through photobiomodulation, thereby promoting the proliferation and collagen secretion of chondrocytes to improve cartilage reshaping and stability.

LncRNA CARD8-AS1 suppresses lung adenocarcinoma progression by enhancing TRIM25-mediated ubiquitination of TXNRD1.

Long non-coding RNAs (lncRNAs) play crucial roles in the tumorigenesis and progression of lung adenocarcinoma (LUAD). However, little was known about the role of lncRNAs in high-risk LUAD subtypes: micropapillary-predominant adenocarcinoma (MPA) and solid-predominant adenocarcinoma (SPA). In this study, we conducted a systematic screening of differentially expressed lncRNAs using RNA sequencing in 10 paired MPA/SPA tumor tissues and adjacent normal tissues. Consequently, 110 significantly up-regulated lncRNAs and 288 aberrantly down-regulated lncRNAs were identified (|Log2 Foldchange| ≥ 1 and corrected P < 0.05). The top 10 lncRNAs were further analyzed in 89 MPA/SPA tumor tissues and 59 normal tissues from The Cancer Genome Atlas database. Among them, CARD8-AS1 showed the most significant differential expression, and decreased expression of CARD8-AS1 was significantly associated with a poorer prognosis. Functionally, CARD8-AS1 overexpression remarkably suppressed the proliferation, migration and invasion of LUAD cells both in vitro and in vivo. Conversely, inhibition of CARD8-AS1 yielded opposite effects. Mechanistically, CARD8-AS1 acted as a scaffold to facilitate the interaction between TXNRD1 and E3 ubiquitin ligase TRIM25, thereby promoting the degradation of TXNRD1 through the ubiquitin-proteasome pathway. Additionally, TXNRD1 was found to promote LUAD cell proliferation, migration and invasion in vitro. Furthermore, the suppressed progression of LUAD cells resulting from CARD8-AS1 overexpression could be significantly reversed by simultaneous overexpression of TXNRD1. In conclusion, this study revealed that the lncRNA CARD8-AS1 played a suppressive role in the progression of LUAD by enhancing TRIM25-mediated ubiquitination of TXNRD1. The CARD8-AS1-TRIM25-TXNRD1 axis may represent a promising therapeutic target for LUAD.

Metabolic Abnormalities Linked to Auditory Pathways in ApoE-Knockout HEI-OC1 Cells: A Transcription-Metabolism Co-Analysis.

Lipid transporter protein apolipoprotein E (APOE) has contributed to functional studies of various organ functions. Animals with ApoE knockout (KO) have been used to study atherosclerosis and hyperlipidemia while an increasing number of researchers have recently focused on the association of ApoE with hearing loss. A study found that ApoE KO mice experience sensorineural hearing loss and hair cell loss, but the exact mechanism is unclear. To explore the potential relationship between ApoE and hearing loss, we used HEI-OC1 cells (House Ear Institute-Organ of Corti) with Corti apparatus properties to reveal cell changes after ApoE knockout by combined transcriptome and metabolomic analysis. We found that glutamate deficiency, caused by reduced expression of glutamine transporter proteins, was a key correlate of basal metabolism and that inadequate glutamate causes apoptosis by reducing the cells' resistance to external damage. Our study provides a reference mechanism for hearing loss due to ApoE KO.

NPC1 Deficiency Contributes to Autophagy-Dependent Ferritinophagy in HEI-OC1 Auditory Cells.

Niemann-Pick type C disease (NPCD) is a rare genetic syndrome characterized by cholesterol accumulation in multiple organelles. NPCD is mainly caused by gene deficiency of NPC intracellular cholesterol transporter 1 (NPC1). It has been reported that some of the NPCD patients exhibit clinical features of progressive hearing loss at high frequency and iron disorder, but the underlying relationship is unknown. A recent study has reported that ferroptosis contributes to the impairment of cochlear hair cells that are related to sensory hearing. In this study, we generated NPC1-deficient HEI-OC1 cells to show the effect of NPC1 deficiency on cochlear outer hair cells. We found that NPC1 deficiency enhances autophagy-dependent ferritinophagy to release Fe (II). Our work provides important insights into the effect of NPC1 deficiency in auditory cells, indicating that it induces ferroptosis and results in hearing loss.

Mutations in OSBPL2 cause hearing loss associated with primary cilia defects via sonic hedgehog signaling.

Defective primary cilia cause a range of diseases called ciliopathies, which include hearing loss (HL). Variants in the human oxysterol-binding protein like 2 (OSBPL2/ORP2) are responsible for autosomal dominant nonsyndromic HL (DFNA67). However, the pathogenesis of OSBPL2 deficiency has not been fully elucidated. In this study, we show that the Osbpl2-KO mice exhibited progressive HL and abnormal cochlear development with defective cilia. Further research revealed that OSBPL2 was located at the base of the kinocilia in hair cells (HCs) and primary cilia in supporting cells (SCs) and functioned in the maintenance of ciliogenesis by regulating the homeostasis of PI(4,5)P2 (phosphatidylinositol 4,5-bisphosphate) on the cilia membrane. OSBPL2 deficiency led to a significant increase of PI(4,5)P2 on the cilia membrane, which could be partially rescued by the overexpression of INPP5E. In addition, smoothened and GL13, the key molecules in the Sonic Hedgehog (Shh) signaling pathway, were detected to be downregulated in Osbpl2-KO HEI-OC1 cells. Our findings revealed that OSBPL2 deficiency resulted in ciliary defects and abnormal Shh signaling transduction in auditory cells, which helped to elucidate the underlying mechanism of OSBPL2 deficiency in HL.

Oxysterol-binding protein-like 2 contributes to the developmental progression of preadipocytes by binding to ß-catenin.

Oxysterol-binding protein-like 2 (OSBPL2), also known as oxysterol-binding protein-related protein (ORP) 2, is a member of lipid transfer protein well-known for its role in regulating cholesterol homeostasis. A recent study reported that OSBPL2/ORP2 localizes to lipid droplets (LDs) and is associated with energy metabolism and obesity. However, the function of OSBPL2/ORP2 in adipocyte differentiation is poorly understood. Here, we report that OSBPL2/ORP2 contributes to the developmental progression of preadipocytes. We found that OSBPL2/ORP2 binds to ß-catenin, a key effector in the Wnt signaling pathway that inhibits adipogenesis. This complex plays a role in regulating the protein level of ß-catenin only in preadipocytes, not in mature adipocytes. Our data further indicated that OSBPL2/ORP2 mediates the transport of ß-catenin into the nucleus and thus regulates target genes related to adipocyte differentiation. Deletion of OSBPL2/ORP2 markedly reduces ß-catenin both in the cytoplasm and in the nucleus, promotes preadipocytes maturation, and ultimately leads to obesity-related characteristics. Altogether, we provide novel insight into the function of OSBPL2/ORP2 in the developmental progression of preadipocytes and suggest OSBPL2/ORP2 may be a potential therapeutic target for the treatment of obesity-related diseases.

A Retrospective Study on the Use of Chinese Patent Medicine in 24 Medical Institutions for COVID-19 in China.

Objective: This research aims to analyze the application regularity of Chinese patent medicine during the COVID-19 epidemic by collecting the names of the top three Chinese patent medicines used by 24 hospitals in 14 provinces of China in four time periods (January 20-22, February 16-18, March 01-03, April 01-03, 2020), and explore its contribution to combating the disease. Methods: 1) We built a database of the top three Chinese patent medicines used by 24 hospitals. 2) The frequency and efficacy distribution of Chinese patent medicine were analyzed with risk areas, regions, and hospitals of different properties as three factors. 3) Finally, we analyzed the differences in the use of heat-clearing and non-heat-clearing medicines among the three factors (χ2 test) and the correlation between the Chinese patent medicine and COVID-19 epidemic (correlation analysis) with SPSS 23.0 statistical software. Results: 1) The heat-clearing medicine was the main use category nationwide during January 20-22, 2020. Meanwhile, there was a significant difference in the utilization rate of heat-clearing and non-heat-clearing medicine in different risk areas (p < 0.01). 2) The variety of Chinese patent medicine was increased nationwide during February 16-18, 2020, mainly including tonics, blood-activating and resolving-stasis, and heat-clearing medicines. Meanwhile, there was a significant difference in the utilization rate of heat-clearing and non-heat-clearing medicine in the southern and northern regions (p < 0.05). 3) Tonics, and blood-activating and resolving-stasis medicines became the primary use categories nationwide during March 01-03, 2020. 4) The tonics class, and blood-activating and resolving-stasis medicine were still the primary categories nationwide during April 01-03, 2020. Meanwhile, there was a significant difference in the utilization rate of heat-clearing and non-heat-clearing medicine in different risk areas (p < 0.01). Conclusion: Chinese patent medicine has a certain degree of participation in fighting against the COVID-19. The efficacy distribution is related to the risk area, region, and hospital of different properties, among which the risk area is the main influencing factor. It is hoped that future research can further collect the application amount of Chinese patent medicine used in hospitals all over the country, so as to perfectly reflect the relationship between Chinese patent medicine and the epidemic situation.

Comparative transcriptome analysis of auditory OC-1 cells and zebrafish inner ear tissues in the absence of human OSBPL2 orthologues.

In our previous study, Oxysterol-binding protein-related protein 2 (OSBPL2) was first identified as a new deafness-causative gene contribute to non-syndromic hearing loss. However, the underlying mechanism of OSBPL2-induced hearing loss remains unknown. Here, we used hearing-specific cells and tissues OC-1 cells and zebrafish inner ear tissues as models to identify common transcriptome changes in genes and pathways in the absence of human OSBPL2 orthologues by RNA-seq analysis. In total, 2112 differentially expressed genes (DEGs) were identified between wild-type (WT) and Osbpl2-/- OC-1 cells, and 877 DEGs were identified between WT and osbpl2b-/- zebrafish inner ear tissues. Functional annotation implicated Osbpl2/osbpl2b in lipid metabolism, cell adhesion and the extracellular matrix in both OC-1 cells and zebrafish inner ear tissues. Protein-protein interaction (PPI) analysis indicated that Osbpl2/osbpl2b were also involved in ubiquitination. Further experiments showed that Osbpl2-/- OC-1 cells exhibited an abnormal focal adhesion morphology characterized by inhibited FAK activity and impaired cell adhesion. In conclusion, we identified novel pathways modulated by OSBPL2 orthologues, providing new insight into the mechanism of hearing loss induced by OSBPL2 deficiency.

Deletion of OSBPL2 in auditory cells increases cholesterol biosynthesis and drives reactive oxygen species production by inhibiting AMPK activity.

Oxysterol-binding protein like 2 (OSBPL2) was identified as a novel causal gene for autosomal dominant nonsyndromic hearing loss. However, the pathogenesis of OSBPL2 deficits in ADNSHL was still unclear. The function of OSBPL2 as a lipid-sensing regulator in multiple cellular processes suggested that OSBPL2 might play an important role in the regulation of cholesterol-homeostasis, which was essential for inner ear. In this study the potential roles of OSBPL2 in cholesterol biosynthesis and ROS production were investigated in Osbpl2-KO OC1 cells and osbpl2b-KO zebrafish. RNA-seq-based analysis suggested that OSBPL2 was implicated in cholesterol biosynthesis and AMPK signaling pathway. Furthermore, Osbpl2/osbpl2b-KO resulted in a reduction of AMPK activity and up-regulation of Srebp2/srebp2, Hmgcr/hmgcr and Hmgcs1/hmgcs1, key genes in the sterol biosynthetic pathway and associated with AMPK signaling. In addition, OSBPL2 was also found to interact with ATIC, key activator of AMPK. The levels of total cholesterol and ROS in OC1 cells or zebrafish inner ear were both increased in Osbpl2/osbpl2b-KO mutants and the mitochondrial damage was detected in Osbpl2-KO OC1 cells. This study uncovered the regulatory roles of OSBPL2 in cellular cholesterol biosynthesis and ROS production. These founds might contribute to the deep understanding of the pathogenesis of OSBPL2 mutation in ADNSHL.

OSBPL2 deficiency upregulate SQLE expression increasing intracellular cholesterol and cholesteryl ester by AMPK/SP1 and SREBF2 signalling pathway.

Previous studies have shown that oxysterol binding protein like 2 (OSBPL2) knockdown is closely related to cholesterol metabolism. However, whether there is a direct relation between OSBPL2 and cholesterol synthesis is unknown. This study explored the mechanism of OSBPL2 deficiency in the upregulation of squalene epoxidase (SQLE) and the subsequent accumulation of intracellular cholesterol and cholesteryl ester. Here, we constructed an OSBPL2-deleted HeLa cell line using CRISPR/Cas9 technology, screened differentially expressed genes and examined the transcriptional regulation of SQLE using a dual-luciferase reporter gene. RNA-seq analysis showed that SQLE was upregulated significantly and the dual luciferase reporter gene assay revealed that two new functional transcription factor binding sites of Sp1 transcription factor (SP1) and sterol regulatory element-binding transcription factor 2 (SREBF2) in the SQLE promoter participated in the SQLE transcription and expression. In addition, we also observed that OSBPL2 deletion inhibited the AMPK signalling pathway and that the inhibition of AMPK signalling promoted SP1 and SREBF2 entry into the nuclear to upregulate SQLE expression. Therefore, these data support that OSBPL2 deficiency upregulates SQLE expression and increases the accumulation of cholesterol and cholesteryl ester by suppressing AMPK signalling, which provides new evidence of the connection between OSBPL2 and cholesterol synthesis.

25-hydroxycholesterol down-regulates oxysterol binding protein like 2 (OSBPL2) via the p53/SREBF2/NFYA signaling pathway.

Oxysterol Binding Protein Like 2 (OSBPL2) is a lipid-binding protein implicated in various cellular processes. Previous studies have shown that depression of OSBPL2 significantly increases the level of cellular 25-hydroxycholesterol (25-OHC) which regulates the expression of lipid-metabolism-related genes. However, whether 25-OHC can regulate the expression of OSBPL2 remains unanswered. This study aimed to explore the molecular mechanism of 25-OHC regulating the expression of OSBPL2. Using dual-luciferase reporter assay, we found a decrease of nuclear transcription factor Y subunit alpha (NFYA) bound with OSBPL2 promoter when HeLa cells were treated with 25-OHC. Furthermore, transcriptome sequencing and RNA interference results revealed that the p53/sterol regulatory element binding transcription factor 2 (SREBF2) signaling pathway was involved in the NFYA-dependent transcription of OSBPL2 induced by 25-OHC. Based on these results, we concluded that pleomorphic adenoma gene 1 (PLAG1) and NFYA participated in the basal transcription of OSBPL2 and that 25-OHC decreased the transcription of OSBPL2 via the p53/SREBF2/NFYA signaling pathway. 25-OHC will accumulate over time in OSBPL2 knockdown cells. These results may provide a new insight into the deafness caused by OSBPL2 mutation.

Transfection of IL-2 and/or IL-12 genes into spleen in treatment of rat liver cancer.

AIM: To test the efficacy of gene therapy in rat liver tumor. METHODS: A retroviral vector GCIL12EIL2PN encoding human IL-2 (hIL-2) and mouse IL-12 (mIL-12) fused gene and its packaging cell were constructed. The packaging cell lines contained of IL-2 and/or IL-12 genes were injected intrasplenically to transfect splenocyte at different time. The therapeutic effect, immune function and toxic effect were evaluated. RESULTS: The average survival times of the 4 groups using IL genes at days 1, 3, 5 and 7 after tumor implantation were 53.3+/-3.7, 49.3+/-4.2, 31.0+/-2.1 and 24.3+/-1.4 d respectively in IL-2/IL-12 fused gene group, 25.0+/-2.5, 23.5+/-2.0, 18.3+/-2.4 and 12.0+/-1.8 d respectively in IL-2 gene treatment group, and 39.0+/-4.8, 32.0+/-3.9, 23.0+/-2.5 and 19.4+/-2.1 d respectively in IL-12 gene treatment group (P<0.01, n=10). In the IL-12/IL-2 fused gene treatment group, 30% of rats treated at days 1 and 3 survived more than 60 d and serum mIL-12 and hIL-2 levels were still high at day 3 after treatment. Compared with IL alone, NK cell activity was strongly stimulated by IL-2/IL-12 gene. Microscopy showed that livers were infiltrated by a number of lymphocytes. CONCLUSION: IL-2 and/or IL-12 genes injected directly into spleen increase serum IL-2 and IL-12 levels and enhance the NK cell activity, which may inhibit the liver tumor growth. The therapy of fused gene IL-2/IL-12 is of low toxicity and relatively high NK cell activity. Our data suggest that IL-2/IL-12 fused gene may be a safe and efficient gene therapy for liver tumor. The gene therapy should be administrated as early as possible.