LOXL1-AS1 contributes to metastasis in sonic-hedgehog medulloblastoma by promoting cancer stem-like phenotypes.

BACKGROUND: Medulloblastomas (MBs) are one of the most common malignant brain tumor types in children. MB prognosis, despite improvement in recent years, still depends on clinical and biological risk factors. Metastasis is the leading cause of MB-related deaths, which highlights an unmet need for risk stratification and targeted therapy to improve clinical outcomes. Among the four molecular subgroups, sonic-hedgehog (SHH)-MB harbors clinical and genetic heterogeneity with a subset of high-risk cases. Recently, long non-coding (lnc)RNAs were implied to contribute to cancer malignant progression, but their role in MB remains unclear. This study aimed to identify pro-malignant lncRNAs that have prognostic and therapeutic significance in SHH-MB. METHODS: The Daoy SHH-MB cell line was engineered for ectopic expression of MYCN, a genetic signature of SHH-MB. MYCN-associated lncRNA genes were identified using RNA-sequencing data and were validated in SHH-MB cell lines, MB tissue samples, and patient cohort datasets. SHH-MB cells with genetic manipulation of the candidate lncRNA were evaluated for metastatic phenotypes in vitro, including cell migration, invasion, sphere formation, and expressions of stemness markers. An orthotopic xenograft mouse model was used to evaluate metastasis occurrence and survival. Finally, bioinformatic screening and in vitro assays were performed to explore downstream mechanisms. RESULTS: Elevated lncRNA LOXL1-AS1 expression was identified in MYCN-expressing Daoy cells and MYCN-amplified SHH-MB tumors, and was significantly associated with lower survival in SHH-MB patients. Functionally, LOXL1-AS1 promoted SHH-MB cell migration and cancer stemness in vitro. In mice, MYCN-expressing Daoy cells exhibited a high metastatic rate and adverse effects on survival, both of which were suppressed under LOLX1-AS1 perturbation. Integrative bioinformatic analyses revealed associations of LOXL1-AS1 with processes of cancer stemness, cell differentiation, and the epithelial-mesenchymal transition. LOXL1-AS1 positively regulated the expression of transforming growth factor (TGF)-ß2. Knockdown of TGF-ß2 in SHH-MB cells significantly abrogated their LOXL1-AS1-mediated prometastatic functions. CONCLUSIONS: This study proved the functional significance of LOXL1-AS1 in SHH-MB metastasis by its promotion of TGF-ß2-mediated cancer stem-like phenotypes, providing both prognostic and therapeutic potentials for targeting SHH-MB metastasis.

Role of Peritoneal Equilibration Test in Assessing Folate Transport During Peritoneal Dialysis.

OBJECTIVES: Low plasma folate levels have been reported in patients undergoing hemodialysis and peritoneal dialysis (PD) in clinical studies. However, folate transport has never been mentioned as a factor contributing to low plasma folate levels in patients undergoing PD. The peritoneal equilibrium test (PET) assesses the plasma creatinine level and glucose transport abilities. This study aimed to evaluate the association between plasma folate levels and folate transport during PD based on PET grades. METHODS: This study recruited 50 patients who underwent PD for ≥3 months and were categorized according to PET grades. Data regarding plasma folate levels and dialysate folate were collected. The primary outcomes were the relationship between the PET grade and plasma folate level and between the PET grade and dialysate-to-plasma folate concentration ratio (D/P folate). Furthermore, the difference in the plasma folate level and D/P folate between men and women was assessed. RESULTS: The plasma folate level and the D/P folate significantly differed among the 4 PET groups (both P < .001). PET grade was significantly negatively correlated with plasma folate levels (r = -0.56, P < .001) and positively correlated with D/P folate (r = 0.686, P < .001). In subgroup analysis, neither the plasma folate level nor the D/P folate significantly differed between men and women. CONCLUSIONS: Our study provides clinical evidence that the PET grade is associated with the plasma folate level and D/P folate, regardless of sex. Larger cohort studies are warranted to assess the importance of folate supplementation during PD based on PET grades.

Plant-derived galactolipids enhance specific antibody production and induce class-switch as vaccine adjuvant.

Various plant-derived compounds can activate immune responses against bacterial infections, and this property contributes to them being developed as effective and safe adjuvants for vaccines. This study evaluated the potential adjuvant effects of a galactolipid-enriched fraction generated from the medicinal plant Crassocephalum rabens (designated CRA). Heat shock protein 60 of periodontal disease pathogen Actinobacillus actinomycetemcomitans (AaHSP60) was taken as an antigen and mixed with CRA. The AaHSP60/CRA mixture was then injected intraperitoneally into the BALB/c mice. Titers and affinity of specific antibodies were measured by ELISA. Cytokine profiles in mouse serum or culture media of AaHSP60/CRA-treated splenocytes were analyzed by cytokine multiplex assay and ELISA kits. B cell differentiation and macrophage activation were determined by phenotyping. CRA dramatically enhanced specific antibody titers and induced Ig class switch, as shown by increases in the IgG2a, IgG2b, and IgG3 proportions of total Ig in mouse serum. Furthermore, CRA-induced anti-AaHSP60 antibodies had cross-reactivity to other bacterial HSP60s. Cell-based and animal results demonstrated that CRA induced the release of IL-21 and B cell activating factor (BAFF), which stimulated B cell differentiation. CRA enhanced cell proliferation, uptake ability, and antigen presentation in mouse phagocytes. CRA served as a vaccine adjuvant that enhance mouse immunity against pathogenic antigens. CRA strengthened the activation and capabilities of phagocytes and B cells. Therefore, CRA may be a promising adjuvant for bacterial vaccines including periodontal disease.

Integrated Metabolomic and Transcriptomic Analyses Reveal the Potential Regulation of Flavonoids in the Production of Embryogenic Cultures during Early Somatic Embryogenesis of Longan (Dimocarpus longan Lour.).

Embryogenic cultures of longan (Dimocarpus longan Lour.) contain various metabolites with pharmacological properties that may function in the regulation of somatic embryogenesis (SE). In this study, based on widely targeted metabolomics, 501 metabolites were obtained from the embryogenic calli, incomplete compact proembryogenic cultures, and globular embryos during early SE of longan, among which 41 flavonoids were differentially accumulated during the SE. Using RNA sequencing, 36 flavonoid-biosynthesis-related genes and 43 MYB and 52 bHLH transcription factors were identified as differentially expressed genes. Furthermore, Kyoto Encyclopedia of Genes and Genomes enrichment analysis revealed that the flavonoid metabolism-related pathways were significantly enriched during the early SE. These results suggested that the changes in flavonoid levels in the embryogenic cultures of longan were mediated by MYBs and bHLHs via regulating flavonoid-biosynthesis-related genes, thus potentially regulating early SE. The identified metabolites in the embryogenic cultures of longan can be used to develop pharmaceutical ingredients.

Differential expression of GABAA receptor subunits δ and α6 mediates tonic inhibition in parvalbumin and somatostatin interneurons in the mouse hippocampus.

Inhibitory γ-aminobutyric acid (GABA)-ergic interneurons mediate inhibition in neuronal circuitry and support normal brain function. Consequently, dysregulation of inhibition is implicated in various brain disorders. Parvalbumin (PV) and somatostatin (SST) interneurons, the two major types of GABAergic inhibitory interneurons in the hippocampus, exhibit distinct morpho-physiological properties and coordinate information processing and memory formation. However, the molecular mechanisms underlying the specialized properties of PV and SST interneurons remain unclear. This study aimed to compare the transcriptomic differences between these two classes of interneurons in the hippocampus using the ribosome tagging approach. The results revealed distinct expressions of genes such as voltage-gated ion channels and GABAA receptor subunits between PV and SST interneurons. Gabrd and Gabra6 were identified as contributors to the contrasting tonic GABAergic inhibition observed in PV and SST interneurons. Moreover, some of the differentially expressed genes were associated with schizophrenia and epilepsy. In conclusion, our results provide molecular insights into the distinct roles of PV and SST interneurons in health and disease.

Krüppel-like factor 12 regulates aging ovarian granulosa cell apoptosis by repressing SPHK1 transcription and sphingosine-1-phosphate (S1P) production.

Oxidative stress triggered by aging, radiation, or inflammation impairs ovarian function by inducing granulosa cell (GC) apoptosis. However, the mechanism inducing GC apoptosis has not been characterized. Here, we found that ovarian GCs from aging patients showed increased oxidative stress, enhanced reactive oxygen species activity, and significantly decreased expression of the known antiapoptotic factor sphingosine-1-phosphate/sphingosine kinase 1 (SPHK1) in GCs. Interestingly, the expression of Krüppel-like factor 12 (KLF12) was significantly increased in the ovarian GCs of aging patients. Furthermore, we determined that KLF12 was significantly upregulated in hydrogen peroxide-treated GCs and a 3-nitropropionic acid-induced in vivo model of ovarian oxidative stress. This phenotype was further confirmed to result from inhibition of SPHK1 by KLF12. Interestingly, when endogenous KLF12 was knocked down, it rescued oxidative stress-induced apoptosis. Meanwhile, supplementation with SPHK1 partially reversed oxidative stress-induced apoptosis. However, this function was lost in SPHK1 with deletion of the binding region to the KLF12 promoter. SPHK1 reversed apoptosis caused by hydrogen peroxide-KLF12 overexpression, a result further confirmed in an in vitro ovarian culture model and an in vivo 3-nitropropionic acid-induced ovarian oxidative stress model. Overall, our study reveals that KLF12 is involved in regulating apoptosis induced by oxidative stress in aging ovarian GCs and that sphingosine-1-phosphate/SPHK1 can rescue GC apoptosis by interacting with KLF12 in negative feedback.

Tetra-O-methyl-nordihydroguaiaretic acid inhibits energy metabolism and synergistically induces anticancer effects with temozolomide on LN229 glioblastoma tumors implanted in mice while preventing obesity in normal mice that consume high-fat diets.

Tetra-O-methyl-nordihydroguaiaretic acid (terameprocol; M4N), a global transcription inhibitor, in combination with a second anticancer drug induces strong tumoricidal activity and has the ability to suppress energy metabolism in cultured cancer cells. In this study, we showed that after continuous oral consumption of high-fat (HF) diets containing M4N, the M4N concentration in most of the organs in mice reached ~1 µM (the M4N concentration in intestines and fat pads was as high as 20-40 µM) and treatment with the combination of M4N with temozolomide (TMZ) suppressed glycolysis and the tricarboxylic acid cycle in LN229 human glioblastoma implanted in xenograft mice. Combination treatment of M4N with TMZ also reduced the levels of lactate dehydrogenase A (LDHA), a key enzyme for glycolysis; lactate, a product of LDHA-mediated enzymatic activity; nicotinamide phosphoribosyltransferase, a rate-limiting enzyme for nicotinamide adenine dinucleotide plus hydrogen (NADH)/NAD+ salvage pathway; and NAD+, a redox electron carrier essential for energy metabolism. It was also shown that M4N suppressed oxygen consumption in cultured LN229 cells, indicating that M4N inhibited oxidative phosphorylation. Treatment with M4N and TMZ also decreased the level of hypoxia-inducible factor 1A, a major regulator of LDHA, under hypoxic conditions. The ability of M4N to suppress energy metabolism resulted in induction of the stress-related proteins activating transcription factor 4 and cation transport regulator-like protein 1, and an increase in reactive oxygen species production. In addition, the combination treatment of M4N with TMZ reduced the levels of oncometabolites such as 2-hydroxyglutarate as well as the aforementioned lactate. M4N also induced methylidenesuccinic acid (itaconate), a macrophage-specific metabolite with anti-inflammatory activity, in tumor microenvironments. Meanwhile, the ability of M4N to suppress energy metabolism prevented obesity in mice consuming HF diets, indicating that M4N has beneficial effects on normal tissues. The dual ability of combination treatment with M4N to suppress both energy metabolism and oncometabolites shows that it is potentially an effective therapy for cancer.

Phytogalactolipids activate humoral immunity against colorectal cancer.

BACKGROUND: Colorectal cancer (CRC) is the third most lethal cancer in the world, and its incidence is steadily rising. In this study, we investigated the induction of humoral immunity by a phytogalactolipid enriched fraction (CRA) derived from the medicinal plant Crassocephalum rabens (Benth.) S. Moore to combat CRC. METHODS: Immunocompetent BALB/c mice were used to evaluate CRA's therapeutic effects in CRC. The phenotypes of B cell subsets in splenocytes and tumors from the CRA-treated mice were isolated and analyzed by flow cytometry. The titers, isotypes, specificity, antigen recognition, and cytotoxic activity of CRA-induced anti-tumor antibodies were determined. The mechanisms of CRA on B cell differentiation were determined by cell-based analyses, including co-cultural with T cells, cytokine analysis, gene expression by qPCR, and protein expression by western blotting. RESULTS: CRA efficiently inhibited tumor growth in colorectal tumor-bearing allograft mice. CRA treatment attracted an abundance of B cells into the tumor consequently enhancing the anti-tumor antibodies in sera and inducing a class-switch. CRA-induced antisera (designated CRA antisera) specifically recognized surface antigens on the plasma membrane of cancer cells. CRA antisera induced cytotoxicity including antibody-dependent cell cytotoxicity, phagocytosis, and complement-dependent cytotoxicity. CRA interacted with IL-6 receptor to activate STAT3 and cMaf, resulting in T cell secretion of IL-21, which, in turn induced B cell differentiation through the IL-21R/STAT3/Blimp-1 pathway. CONCLUSIONS: CRA regulated T cell activity resulting in B cell activation and triggering of anti-tumor antibodies to impede CRC progression.

Urinary di-(2-ethylhexyl) phthalate metabolites are independently related to serum neurofilament light chain, a biomarker of neurological diseases, in adults: results from NHANES 2013-2014.

Di (2-ethylhexyl) phthalate (DEHP) is a chemical commonly used in the manufacturing of plastics and can pose human health risks, including endocrine disruption, reproductive toxicity, and potential carcinogenic effects. Children may be particularly vulnerable to the harmful effects of DEHP. Early exposure to DEHP has been linked to potential behavioral and learning problems. However, there are no reports to date on whether DEHP exposure in adulthood has neurotoxic effects. Serum neurofilament light chain (NfL), a protein released into the blood after neuroaxonal damage, has been shown to be a reliable biomarker for many neurological diseases. To date, no study has examined the relationship between DEHP exposure and NfL. For the present study, we selected 619 adults (aged ≥ 20 years) from the 2013-2014 National Health and Nutrition Examination Survey (NHANES) to examine the association between urinary DEHP metabolites and serum NfL. We reported higher urinary levels of ln-mono(2-ethyl-5-hydroxyhexyl) phthalate (MEHHP), ln-mono(2-ethyl-5-oxohexyl) phthalate (MEOHP), and ln-mono(2-ethyl-5-carboxypentyl) phthalate (MECPP), and ln-ΣDEHP levels were associated with higher serum levels of ln-NfL (ΣDEHP: ß-coefficient = 0. 075; S.E. = 0.026; P = 0.011). When we divided ΣDEHP into quartiles, mean NfL concentrations increased with quartiles of MEHHP (P for trend = 0.023). The association was more pronounced in males, non-Hispanic white race, higher income, and BMI < 25. In conclusion, higher DEHP exposure was positively associated with higher serum NfL in adults from NHANES 2013-2014. If this finding is causal, it is possible that DEHP exposure in adulthood may also induce neurological damage. Although the causality of this observation and the clinical significance are uncertain, our findings suggest that additional research is needed on DEHP exposure, serum NfL, and neurological disease in adults.

A flexible liposomal polymer complex as a platform of specific and regulable immune regulation for individual cancer immunotherapy.

BACKGROUND: The applicability and therapeutic efficacy of specific personalized immunotherapy for cancer patients is limited by the genetic diversity of the host or the tumor. Side-effects such as immune-related adverse events (IRAEs) derived from the administration of immunotherapy have also been observed. Therefore, regulatory immunotherapy is required for cancer patients and should be developed. METHODS: The cationic lipo-PEG-PEI complex (LPPC) can stably and irreplaceably adsorb various proteins on its surface without covalent linkage, and the bound proteins maintain their original functions. In this study, LPPC was developed as an immunoregulatory platform for personalized immunotherapy for tumors to address the barriers related to the heterogenetic characteristics of MHC molecules or tumor associated antigens (TAAs) in the patient population. Here, the immune-suppressive and highly metastatic melanoma, B16F10 cells were used to examine the effects of this platform. Adsorption of anti-CD3 antibodies, HLA-A2/peptide, or dendritic cells' membrane proteins (MP) could flexibly provide pan-T-cell responses, specific Th1 responses, or specific Th1 and Th2 responses, depending on the host needs. Furthermore, with regulatory antibodies, the immuno-LPPC complex properly mediated immune responses by adsorbing positive or negative antibodies, such as anti-CD28 or anti-CTLA4 antibodies. RESULTS: The results clearly showed that treatment with LPPC/MP/CD28 complexes activated specific Th1 and Th2 responses, including cytokine release, CTL and prevented T-cell apoptosis. Moreover, LPPC/MP/CD28 complexes could eliminate metastatic B16F10 melanoma cells in the lung more efficiently than LPPC/MP. Interestingly, the melanoma resistance of mice treated with LPPC/MP/CD28 complexes would be reversed to susceptible after administration with LPPC/MP/CTLA4 complexes. NGS data revealed that LPPC/MP/CD28 complexes could enhance the gene expression of cytokine and chemokine pathways to strengthen immune activation than LPPC/MP, and that LPPC/MP/CTLA4 could abolish the LPPC/MP complex-mediated gene expression back to un-treatment. CONCLUSIONS: Overall, we proved a convenient and flexible immunotherapy platform for developing personalized cancer therapy.

HBV genotype-dependent association of HLA variants with the serodecline of HBsAg in chronic hepatitis B patients.

Seroclearance of hepatitis B surface antigen (HBsAg) is regarded as the functional cure for chronic hepatitis B (CHB). The relationship between human leukocyte antigen (HLA) variants, hepatitis B virus genotype, and longitudinal HBsAg serodecline remains to be explored. A total of 1735 HBeAg-seronegative CHB patients with genotype B or C infection of the community-based REVEAL-HBV cohort were genotyped for rs1710 (HLA-G) and rs2770 (HLA-B) using TaqMan assay. Cox proportional hazard regression and generalized linear mixed models were used to analyze the association of HLA genetic variants with the rate of HBsAg seroclearance and longitudinal HBsAg serodecline. Rs1710 G allele was differentially associated with the HBsAg seroclearance in genotype B [aRR (95% CI) = 0.74 (0.56-0.98)] and genotype C [aRR (95%CI) = 1.43 (1.08-1.88)] infection. Rs2770 G allele was associated with HBsAg seroclearance only in genotype B infection [aRR (95% CI) = 0.69 (0.52-0.91)]. The alleles associated with HBsAg seroclearance were significant predictors for the serodecline of HBsAg levels in an HBV genotype-dependent manner (genotype B infection: rs1710, P = 0.013; rs2770, P = 0.0081; genotype C infection: rs1710, P = 0.0452). Our results suggest both spontaneous HBsAg seroclearance and serodecline are modified by the interaction between HLA variants and HBV genotype.

Cellular mechanisms underlying central sensitization in a mouse model of chronic muscle pain.

Chronic pain disorders are often associated with negative emotions, including anxiety and depression. The central nucleus of the amygdala (CeA) has emerged as an integrative hub for nociceptive and affective components during central pain development. Prior adverse injuries are precipitating factors thought to transform nociceptors into a primed state for chronic pain. However, the cellular basis underlying the primed state and the subsequent development of chronic pain remains unknown. Here, we investigated the cellular and synaptic alterations of the CeA in a mouse model of chronic muscle pain. In these mice, local infusion of pregabalin, a clinically approved drug for fibromyalgia and other chronic pain disorders, into the CeA or chemogenetic inactivation of the somatostatin-expressing CeA (CeA-SST) neurons during the priming phase prevented the chronification of pain. Further, electrophysiological recording revealed that the CeA-SST neurons had increased excitatory synaptic drive and enhanced neuronal excitability in the chronic pain states. Finally, either chemogenetic inactivation of the CeA-SST neurons or pharmacological suppression of the nociceptive afferents from the brainstem to the CeA-SST neurons alleviated chronic pain and anxio-depressive symptoms. These data raise the possibility of targeting treatments to CeA-SST neurons to prevent central pain sensitization.

Clinical and Molecular Features in Medulloblastomas Subtypes in Children in a Cohort in Taiwan.

Medulloblastoma (MB) was classified into four molecular subgroups: WNT, SHH, group 3, and group 4. In 2017, 12 subtypes within 4 subgroups and 8 subtypes within non-WNT/non-SHH subgroups according to the differences of clinical features and biology were announced. In this study, we aimed to identify the heterogeneity of molecular features for discovering subtype specific factors linked to diagnosis and prognosis. We retrieved 70 MBs in children to perform RNA sequencing and a DNA methylation array in Taiwan. Integrated with clinical annotations, we achieved classification of 12 subtypes of pediatric MBs in our cohort series with reference to the other reported series. We analyzed the correlation of cell type enrichment in SHH MBs and found that M2 macrophages were enriched in SHH ß, which related to good outcomes of SHH MBs. The high infiltration of M2 macrophages may be an indicator of a favorable prognosis and therapeutic target for SHH MBs. Furthermore, C11orf95-RELA fusion was observed to be associated with recurrence and a poor prognosis. These results will contribute to the establishment of a molecular diagnosis linked to prognostic indicators of relevance and help to promote molecular-based risk stratified treatment for MBs in children.

[Clinical Characteristics of Nontuberculous Mycobacterium Infection Cases in Sichuan, China in 2016-2021: A Retrospective Study].

Objective: To study the distribution of nontuberculous mycobacterium (NTM) strains, clinical characteristics and drug sensitivity data of NTM infections so as to provide support for the prevention and treatment of diseases caused by NTM infection in Sichuan. Methods: The clinical data of NTM infection cases treated at the Public Health Clinical Center of Chengdu between July 2016 and July 2021 were collected and the characteristics of the infections were retrospectively reviewed. Results: There were differences in sex, age and underlying diseases among the NTM infection cases in Sichuan. Specifically, young and middle-aged men aged between 20 and 40 were susceptible to AIDS, older men aged over 60 were susceptible to lung diseases, and middle-aged and older women over 40 were susceptible to bronchiectasis. Respiratory tract was the main route of NTM infection. The dominant strain in Sichuan was M. chelonae/ abscessus. The drug resistance rate of M. avium and M. chelonae/ abscessus were relatively higher. Conclusion: For NTM infection patients with different demographic characteristics and underlying diseases, the NTM infection sites, strains, and drug resistance are also different. Definite etiological diagnosis is essential to the treatment of NTM infection. We should highlight the importance of adopting individualized treatment for different NTM infections.

In Vivo Evaluation of the Effects of Sintering Temperature on the Optical Properties of Dental Glass-Ceramics.

In prosthodontics, the ability of glass-ceramics to express the optical properties of natural teeth is an important goal of esthetic restorations. Dental restorations do not merely need to be similar in color to natural teeth; proper optical properties, such as opalescence, transparency, etc., must be combined in order to achieve excellent esthetic effects. The optical properties of ceramic materials are mainly distinguished by different hues (e.g., A, B, C, and D) combined with translucency (e.g., high translucency (HT), medium translucency (MT), low translucency (LT), and medium opacity (MO)). However, there are many varieties of tooth color. Therefore, it is expected that glass-ceramics can change their nanocrystal size and porosity through different heat-treatment temperatures and times and, thereby, present different transparency effects. This study mainly analyzed the influence of changes in sintering temperature on the optical properties of glass-ceramics. The optical properties of glass-ceramics in the oral cavity were evaluated with human trials. We hypothesized that (1) the transparency of glass-ceramics can be changed by controlling the sintering temperature and (2) glass-ceramics modified by the sintering temperature can be suitable for clinical applications. Results showed that the transparency decreased, the nanoparticle size increased, the crystallinity increased, and the surface hardness decreased as the sintering temperature increased. High-brightness glass-ceramics have more-sensitive optical properties. Results of clinical trials showed that glass-ceramics whose transparency was changed by controlling the sintering temperature can be candidates for clinical applications. Based on the above results, the hypotheses of this study were supported. In the future, we will continue to explore the esthetic field of dental restorations.

Genome-wide identification of miRNAs and targets associated with cell wall biosynthesis: Differential roles of dlo-miR397a and dlo-miR408-3p during early somatic embryogenesis in longan.

The dynamic alterations in cell wall (CW) biosynthesis play an essential role in physiological isolation during the plant somatic embryogenesis (SE). However, the mechanisms underlying the functions of cell wall-associated miRNAs (CW-miRNA) remain poorly understood in plant SE. Here, we have identified 36 distinct candidate miRNAs associated with CW biosynthesis from longan third-generation genome as well as miRNA transcriptome, and modified RLM-RACE validated four distinct miRNA, which specifically targeted four CW-related genes. More importantly, we found that the dlo-miR397a-antagomir significantly enhanced DlLAC7 expression and improved laccase activity. Interestingly, inhibition of dlo-miR397a increased CW lignin deposition and promoted the tightening of protodermal cell by miRNA-mimic technology during early SE. Moreover, overexpression of dlo-miR408-3p (dlo-miR408-3p-agomir) markedly decreased DlLAC12 expression. dlo-miR408-3p-agomir activated rapid cell division, thus promoting the globular embryo (GE) development, which might be due to high DNA synthesis activity in protoepidermal cells, rather than affecting lignin synthesis. The subcellular location also indicated that both DlLAC7 and DlLAC12 proteins were primarily localized in CW and regulated CW biosynthesis. Overall, our findings provided new insight on the molecular regulatory networks comprising various miRNAs associated with cell wall, and established that dlo-miR397a and dlo-miR408-3p played differential roles during early SE in longan. The findings also shed some light on the potential role of miRNA target DlLAC regulating in vivo embryonic development of plant.

Enrichment of Tumor-Infiltrating B Cells in Group 4 Medulloblastoma in Children.

Medulloblastoma (MB) is the most common malignant brain tumor in children. It is classified into core molecular subgroups (wingless activated (WNT), sonic hedgehog activated (SHH), Group 3 (G3), and Group 4 (G4)). In this study, we analyzed the tumor-infiltrating immune cells and cytokine profiles of 70 MB patients in Taiwan using transcriptome data. In parallel, immune cell composition in tumors from the SickKids cohort dataset was also analyzed to confirm the findings. The clinical cohort data showed the WNT and G4 MB patients had lower recurrence rates and better 5-year relapse-free survival (RFP) compared with the SHH and G3 MB patients, among the four subgroups of MB. We found tumor-infiltrating B cells (TIL-Bs) enriched in the G4 subgroups in the Taiwanese MB patients and the SickKids cohort dataset. In the G4 subgroups, the patients with a high level of TIL-Bs had better 5-year overall survival. Mast cells presented in G4 MB tumors were positively correlated with TIL-Bs. Higher levels of CXCL13, IL-36γ, and CCL27 were found compared to other subgroups or normal brains. These three cytokines, B cells and mast cells contributed to the unique immune microenvironment in G4 MB tumors. Therefore, B-cell enrichment is a G4-subgroup-specific immune signature and the presence of B cells may be an indicator of a better prognosis in G4 MB patients.

Sexual Crossing, Chromosome-Level Genome Sequences, and Comparative Genomic Analyses for the Medicinal Mushroom Taiwanofungus Camphoratus (Syn. Antrodia Cinnamomea, Antrodia Camphorata).

Taiwanofungus camphoratus mushrooms are a complementary and alternative medicine for hangovers, cancer, hypertension, obesity, diabetes, and inflammation. Though Taiwanofungus camphoratus has attracted considerable biotechnological and pharmacological attention, neither classical genetic nor genomic approaches have been properly established for it. We isolated four sexually competent monokaryons from two T. camphoratus dikaryons used for the commercial cultivation of orange-red (HC1) and milky-white (SN1) mushrooms, respectively. We also sequenced, annotated, and comparatively analyzed high-quality and chromosome-level genome sequences of these four monokaryons. These genomic resources represent a valuable basis for understanding the biology, evolution, and secondary metabolite biosynthesis of this economically important mushrooms. We demonstrate that T. camphoratus has a tetrapolar mating system and that HC1 and SN1 represent two intraspecies isolates displaying karyotypic variation. Compared with several edible mushroom model organisms, T. camphoratus underwent a significant contraction in the gene family and individual gene numbers, most notably for plant, fungal, and bacterial cell-wall-degrading enzymes, explaining why T. camphoratus mushrooms are rare in natural environments, are difficult and time-consuming to artificially cultivate, and are susceptible to fungal and bacterial infections. Our results lay the foundation for an in-depth T. camphoratus study, including precise genetic manipulation, improvements to mushroom fruiting, and synthetic biology applications for producing natural medicinal products. IMPORTANCETaiwanofungus camphoratus (Tc) is a basidiomycete fungus that causes brown heart rot of the aromatic tree Cinnamomum kanehirae. The Tc fruiting bodies have been used to treat hangovers, abdominal pain, diarrhea, hypertension, and other diseases first by aboriginal Taiwanese and later by people in many countries. To establish classical genetic and genomic approaches for this economically important medicinal mushroom, we first isolated and characterized four sexually competent monokaryons from two dikaryons wildly used for commercial production of Tc mushrooms. We applied PacBio single molecule, real-time sequencing technology to determine the near-completed genome sequences of four monokaryons. These telomere-to-telomere and gapless haploid genome sequences reveal all genomic variants needed to be studied and discovered, including centromeres, telomeres, retrotransposons, mating type loci, biosynthetic, and metabolic gene clusters. Substantial interspecies diversities are also discovered between Tc and several other mushroom model organisms, including Agrocybe aegerita, Coprinopsis cinerea, and Schizophyllum commune, and Ganoderma lucidum.

Canine transmissible venereal tumour established in immunodeficient mice reprograms the gene expression profiles associated with a favourable tumour microenvironment to enable cancer malignancy.

BACKGROUND: Canine transmissible venereal tumours (CTVTs) can cross the major histocompatibility complex barrier to spread among dogs. In addition to the transmissibility within canids, CTVTs are also known as a suitable model for investigating the tumour-host immunity interaction because dogs live with humans and experience the same environmental risk factors for tumourigenesis. Moreover, outbred dogs are more appropriate than inbred mice models for simulating the diversity of human cancer development. This study built a new model of CTVTs, known as MCTVTs, to further probe the shaping effects of immune stress on tumour development. For xenotransplantation, CTVTs were first injected and developed in immunodeficient mice (NOD.CB17-Prkdcscid/NcrCrl), defined as XCTVTs. The XCTVTs harvested from NOD/SCID mice were then inoculated and grown in beagles and named mouse xenotransplantation of CTVTs (MCTVTs). RESULTS: After the inoculation of CTVTs and MCTVTs into immune-competent beagle dogs separately, MCTVTs grew faster and metastasized more frequently than CTVTs did. Gene expression profiles in CTVTs and MCTVTs were analysed by cDNA microarray to reveal that MCTVTs expressed many tumour-promoting genes involved in chronic inflammation, chemotaxis, extracellular space modification, NF-kappa B pathways, and focal adhesion. Furthermore, several well-known tumour-associated biomarkers which could predict tumour progression were overexpressed in MCTVTs. CONCLUSIONS: This study demonstrated that defective host immunity can result in gene instability and enable transcriptome reprogramming within tumour cells. Fast tumour growth in beagle dogs and overexpression of tumour-associated biomarkers were found in a CTVT strain previously established in immunodeficient mice. In addition, dysregulated interaction of chronic inflammation, chemotaxis, and extracellular space modification were revealed to imply the possibly exacerbating mechanisms in the microenvironments of these tumours. In summary, this study offers a potential method to facilitate tumour progression and provide a niche for discovering tumour-associated biomarkers in cancer research.

Hypoglycemic peptide-enriched hydrolysates of Corbicula fluminea and Chlorella sorokiniana possess synergistic hypoglycemic activity through inhibiting α-glucosidase and dipeptidyl peptidase-4 activity.

BACKGROUND: The prevalence of diabetes mellitus worldwide has increased in recent decades. Maintaining the level of blood glucose is the most basic and important issue for diabetics. This study aimed to investigate the hypoglycemic activity of a combination of hypoglycemic peptide-enriched hydrolysates of Corbicula fluminea (ACH) and Chlorella sorokiniana (PCH). RESULTS: Combined supplementation of ACH and PCH synergistically inhibited α-glucosidase and DPP4 activities in vitro. After 4 weeks of treatment with ACH and/or PCH, the plasma glucose concentration and insulin, homeostasis model assessment-estimated insulin resistance (HOMA-IR), total cholesterol (TC) and triglyceride (TG) levels significantly decreased. The hypoglycemic peptides in ACH and PCH were purified and assayed for α-glucosidase and DPP4 activity. The hypoglycemic peptides in ACH and PCH effectively decreased α-glucosidase and DPP4 activities. In silico assays showed that these two peptide types have different docking poses, which determined their inhibitory effect against α-glucosidase and DPP4 activity. CONCLUSION: Combined treatment with hypoglycemic peptide-enriched ACH and PCH could modulate blood glucose by synergistically inhibiting α-glucosidase and DPP4 activities. © 2021 Society of Chemical Industry.