E3 ubiquitin ligase RNF128 attenuates colitis and colorectal tumorigenesis by triggering the degradation of IL-6 receptors.

INTRODUCTION: Intestinal immune dysregulation is strongly linked to the occurrence and formation of tumors. RING finger protein 128 (RNF128) has been identified to play distinct immunoregulatory functions in innate and adaptive systems. However, the physiological roles of RNF128 in intestinal inflammatory conditions such as colitis and colorectal cancer (CRC) remain controversial. OBJECTIVES: To elucidate the function and mechanism of RNF128 in colitis and CRC. METHODS: Animal models of dextran sodium sulfate (DSS)-induced colitis and azoxymethane (AOM)/DSS-induced CRC were established in WT and Rnf128-deficient mice and evaluated by histopathology. Co-immunoprecipitation and ubiquitination analyses were employed to investigate the role of RNF128 in IL-6-STAT3 signaling. RESULTS: RNF128 was significantly downregulated in clinical CRC tissues compared with paired peritumoral tissues. Rnf128-deficient mice were hypersusceptible to both colitis induced by DSS and CRC induced by AOM/DSS or APC mutation. Loss of RNF128 promoted the proliferation of CRC cells and STAT3 activation during the early transformative stage of carcinogenesis in vivo and in vitro when stimulated by IL-6. Mechanistically, RNF128 interacted with the IL-6 receptor α subunit (IL-6Rα) and membrane glycoprotein gp130 and mediated their lysosomal degradation in ligase activity-dependent manner. Through a series of point mutations in the IL-6 receptor, we identified that RNF128 promoted K48-linked polyubiquitination of IL-6Rα at K398/K401 and gp130 at K718/K816/K866. Additionally, blocking STAT3 activation effectively eradicated the inflammatory damage of Rnf128-deficient mice during the transformative stage of carcinogenesis. CONCLUSION: RNF128 attenuates colitis and colorectal tumorigenesis by inhibiting IL-6-STAT3 signaling, which sheds novel insights into the modulation of IL-6 receptors and the inflammation-to-cancer transition.

AIM2 enhances Candida albicans infection through promoting macrophage apoptosis via AKT signaling.

Candida albicans is among the most prevalent invasive fungal pathogens for immunocompromised individuals and novel therapeutic approaches that involve immune response modulation are imperative. Absent in melanoma 2 (AIM2), a pattern recognition receptor for DNA sensing, is well recognized for its involvement in inflammasome formation and its crucial role in safeguarding the host against various pathogenic infections. However, the role of AIM2 in host defense against C. albicans infection remains uncertain. This study reveals that the gene expression of AIM2 is induced in human and mouse innate immune cells or tissues after C. albicans infection. Furthermore, compared to their wild-type (WT) counterparts, Aim2-/- mice surprisingly exhibit resistance to C. albicans infection, along with reduced inflammation in the kidneys post-infection. The resistance of Aim2-/- mice to C. albicans infection is not reliant on inflammasome or type I interferon production. Instead, Aim2-/- mice display lower levels of apoptosis in kidney tissues following infection than WT mice. The deficiency of AIM2 in macrophages, but not in dendritic cells, results in a phenocopy of the resistance observed in Aim2-/- mice against C. albican infection. The treatment of Clodronate Liposome, a reagent that depletes macrophages, also shows the critical role of macrophages in host defense against C. albican infection in Aim2-/- mice. Furthermore, the reduction in apoptosis is observed in Aim2-/- mouse macrophages following infection or treatment of DNA from C. albicans in comparison with controls. Additionally, higher levels of AKT activation are observed in Aim2-/- mice, and treatment with an AKT inhibitor reverses the host resistance to C. albicans infection. The findings collectively demonstrate that AIM2 exerts a negative regulatory effect on AKT activation and enhances macrophage apoptosis, ultimately compromising host defense against C. albicans infection. This suggests that AIM2 and AKT may represent promising therapeutic targets for the management of fungal infections.

IRF7 Exacerbates Candida albicans Infection by Compromising CD209-Mediated Phagocytosis and Autophagy-Mediated Killing in Macrophages.

IFN regulatory factor 7 (IRF7) exerts anti-infective effects by promoting the production of IFNs in various bacterial and viral infections, but its role in highly morbid and fatal Candida albicans infections is unknown. We unexpectedly found that Irf7 gene expression levels were significantly upregulated in tissues or cells after C. albicans infection in humans and mice and that IRF7 actually exacerbates C. albicans infection in mice independent of its classical function in inducing IFNs production. Compared to controls, Irf7-/- mice showed stronger phagocytosis of fungus, upregulation of C-type lectin receptor CD209 expression, and enhanced P53-AMPK-mTOR-mediated autophagic signaling in macrophages after C. albicans infection. The administration of the CD209-neutralizing Ab significantly hindered the phagocytosis of Irf7-/- mouse macrophages, whereas the inhibition of p53 or autophagy impaired the killing function of these macrophages. Thus, IRF7 exacerbates C. albicans infection by compromising the phagocytosis and killing capacity of macrophages via regulating CD209 expression and p53-AMPK-mTOR-mediated autophagy, respectively. This finding reveals a novel function of IRF7 independent of its canonical IFNs production and its unexpected role in enhancing fungal infections, thus providing more specific and effective targets for antifungal therapy.

Benchtop Nickel Catalysis Invigorated by Electron-Deficient Diene Ligands.

ConspectusDue to the rarity of precious metals like palladium, nickel catalysis is becoming an increasingly important player in organic synthesis, especially for the formation of bonds with sp3-hybridized carbon centers. Traditionally, catalytic processes involving active Ni(0) species have relied on Ni(COD)2 or in situ reduction of Ni(II) salts. However, Ni(COD)2 is an air- and temperature-sensitive material that requires use in an inert-atmosphere glovebox, and in situ reduction protocols of Ni(II) salts using metallic or organometallic reductants add additional complications to reaction development.This Account chronicles the development of air-stable Ni(0) precursors as replacements for Ni(COD)2 or in situ reduction. Based on Schrauzer's seminal discovery of Ni(COD)(DQ) as an air-stable zerovalent organonickel complex, our research laboratories at Scripps Research and Bristol Myers Squibb have developed a class of precatalysts based on the Ni(COD)(EDD) (EDD = electron-deficient diene) framework, relying on the steric and electronic properties of the supporting diene to render the metal center stable to air, moisture, and even silica gel but reactive to ligand substitution and redox changes.The stable Ni(0) complexes can be accessed through ligand exchange with Ni(COD)2, through reduction of Ni(acac)2 using DIBAL-H, or electrochemically via cathodic reduction of Ni(acac)2 to Ni(COD)2, followed by addition of an EDD ligand in one pot. As a toolkit, the complexes demonstrate reactivity that is equivalent or enhanced compared to Ni(COD)2, catalyzing C-C and C-N cross-couplings, Miyaura borylations, C-H activations, and other transformations. Since the initial report on Ni(COD)(DQ), its reactivity in C(sp2)-CN activation, metallophotoredox, and electric field-induced cross-coupling have also been demonstrated.By incorporating the precatalyst toolkit into reaction discovery campaigns, our laboratories have been able to perform C(sp3)-S(alkyl) couplings and metallonitrenoid carboamination, both of which represent challenging transformations that were inaccessible with traditional phosphine, nitrogen, or electron-deficient olefin ligands. Computational and experimental studies demonstrate how the quinone ligands are hemilabile, adopting η1(O)-bound geometries to relieve steric strain or stabilize transition states and intermediates; redox-active, able to transiently oxidize the metal center; and electron-withdrawing or -donating, depending on metal oxidation state and coordination geometry. These studies show how the ligands enable key steps in catalysis beyond imparting air-stability.Since our report documenting the catalytic activity of Ni(COD)(DQ), many other laboratories have also observed unique reactivity with this precatalyst. Ni(COD)(DQ) was found to offer superior reactivity to Ni(COD)2 in C-N cross coupling to form N,N-diaryl sulfonamides and in preparation of biaryls from aryl halides and benzene through a Ni-mediated, base-assisted homolytic aromatic substitution.

Ligand-Promoted Iron-Catalyzed Nitrene Transfer for the Synthesis of Hydrazines and Triazanes through N-Amidation of Arylamines.

Herein, we report that bulky alkylphosphines such as PtBu3 can switch the roles from actor to spectator ligands to promote the FeCl2 -catalyzed N-amidation reaction of arylamines with dioxazolones, giving hydrazides in high efficiency and chemoselectivity. Mechanistic studies indicated that the phosphine ligands could facilitate the decarboxylation of dioxazolones on the Fe center, and the hydrogen bonding interactions between the arylamines and the ligands on Fe nitrenoid intermediates might play a role in modulating the delicate interplay between the phosphine ligand, arylamine, and acyl nitrene N, favoring N-N coupling over N-P coupling. The new ligand-promoted N-amidation protocols offer a convenient way to access various challenging triazane compounds via double or sequential N-amidation of primary arylamines.

MDA5 Enhances Invasive Candida albicans Infection by Regulating Macrophage Apoptosis and Phagocytosis/Killing Functions.

Candida albicans is a common opportunistic pathogenic fungus. The innate immune system provides the first-line host defense against fungal infection. Innate immune receptors and downstream molecules have been shown to play various roles during fungal infection. The innate immune receptor MDA5, encoded by the gene Ifih1, enhances host resistance against viral and Aspergillus fumigatus infection by inducing the production of interferons (IFNs). However, the role of MDA5 in C. albicans infection is still unclear. Here, we found that the gene expression levels of IFIH1 were significantly increased in innate immune cells after C. albicans stimulation through human bioinformatics analysis or mouse experiments. Through in vivo study, MDA5 was shown to enhance host susceptibility to C. albicans infection independent of IFN production. Instead, MDA5 exerted its influence on macrophages and kidneys by modulating the expression of Noxa, Bcl2, and Bax, thereby promoting apoptosis. Additionally, MDA5 compromised killing capabilities of macrophage by inhibition iNOS expression. The introduction of the apoptosis inducer PAC1 further impaired macrophage functions, mimicking the enhancing effect of MDA5 on C. albicans infection. Furthermore, the administration of macrophage scavengers increased the susceptibility of Ifih1-/- mice to C. albicans. The founding suggests that MDA5 promote host susceptibility to invasive C. albicans by enhancing cell apoptosis and compromising macrophage functions, making MDA5 a target to treat candidiasis.

Amphetamine-induced neurite injury in PC12 cells through inhibiting GAP-43 pathway.

Amphetamine (AMPH) causes the degeneration of dopamine terminals in the central nervous system. The mechanisms for this damage are unclear. We found AMPH reduced level of GAP-43 in the striatum of rats that receives rich dopaminergic terminals. Using PC12 cells as dopaminergic neuronal models, we further found that AMPH inhibited GAP-43 and GAP-43 phosphorylation in PC12 cells. The reduced GAP-43 was correlated with neurite injury of PC12 cells. The PKCß1, an upstream molecule of GAP-43, was also inhibited by AMPH. Phorbol 12-myristate 13-acetate (PMA) as a specific activator of PKC increased levels of PKCß1 and GAP-43, and efficiently prevented neurite degeneration of PC12 cells induced by AMPH. On the other side, enzastuarin, an inhibitor of PKC, decreased levels of PKCß1 and GAP-43, and caused neurite injury of PC12 cells. Together, our results suggest that AMPH induces neurite injury in PC12 cells through inhibiting PKCß1/GAP-43 pathway.

Directed, nickel-catalyzed 1,2-alkylsulfenylation of alkenyl carbonyl compounds.

We report a regioselective, nickel-catalyzed syn-1,2-carbosulfenylation of non-conjugated alkenyl carbonyl compounds with alkyl/arylzinc nucleophiles and tailored N-S electrophiles. This method allows the simultaneous installation of a variety of C(sp3) and S(Ar) (or Se(Ar)) groups onto unactivated alkenes, which complements previously developed 1,2-carbosulfenylation methodology in which only C(sp2) nucleophiles are compatible. A bidentate directing auxiliary controls regioselectivity, promotes high syn-stereoselectivity with a variety of E- and Z-internal alkenes, and enables the use of an array of electrophilic sulfenyl (and seleno) electrophiles. Among compatible electrophiles, those with N-alkyl-benzamide leaving groups were found to be especially effective, as determined through comprehensive structure-reactivity mapping.

Next generation sequencing of RB1gene for the molecular diagnosis of ethnic minority with retinoblastoma in Yunnan.

BACKGROUND: Retinoblastoma is a rare intraocular malignancy and typically initiated by inactivating biallelic mutations of RB1 gene. Each year, ~ 8000 children worldwide are diagnosed for retinoblastoma. In high-income countries, patient survival is over 95% while low-income countries is ~ 30%.If disease is diagnosed early and treated in centers specializing in retinoblastoma, the survival might exceed 95% and many eyes could be safely treated and support a lifetime of good vision. In China, approximate 1100 newly diagnosed cases are expected annually and 28 hospitals covering 25 provinces established centers classified by expertise and resources for better treatment options and follow-up. Comparing with other province of eastern China, Yunnan province is remote geographically. This might result that healthcare staff have low awareness of the role of genetic testing in management and screening in families. METHODS: The patients with retinoblastoma were selected in Yunnan. DNA from blood was used for targeted gene sequencing. Then, an in-house bioinformatics pipeline was done to detect both single nucleotide variants and small insertions/deletions. The pathogenic mutations were identified and further confirmed by conventional methods and cosegregation in families. RESULTS: Using our approach, targeted next generation sequencing was used to detect the mutation of these 12 probands. Bioinformatic predictions showed that nine mutations were found in our study and four were novel pathogenic variants in these nine mutations. CONCLUSIONS: It's the first report to describe RB1 mutations in Yunnan children with retinoblastoma. This study would improve role of genetic testing for management and family screening.

Characteristics of olfactory ensheathing cells and microarray analysis in Tupaia belangeri (Wagner, 1841).

Tree shrews are most closely related to the primates and so possess a number of advantages in experimental studies; they have been used as an animal model in bacterial and virus infection, cancer, endocrine system disease, and certain nervous system diseases. Their olfactory ensheathing cells (OECs) are able to release several cytokines to promote neuronal survival, regeneration and remyelination. The present study used western blot analysis to identify antibody specificity in protein extracts from whole tree shrew brains to identify the specificity of p75 nerve growth factor receptor (NGFR) derived from rabbits (75 kDa). OECs were cultured and isolated, then stained and identified using the antibodies for p75NGFR. To investigate the capacity of OECs to express cytokines and growth factors, microarray technology was used, and the analysis revealed that OECs were able to express 9,821 genes. Of these genes, 44 genes were from the neurotrophic factor family, which may indicate their potential in transplantation in vivo. The present study considered the function of OECs as revealed by other studies, and may contribute to future research.

Atractylenolide II reverses the influence of lncRNA XIST/miR-30a-5p/ROR1 axis on chemo-resistance of colorectal cancer cells.

This investigation was conducted to elucidate whether atractylenolide II could reverse the role of lncRNA XIST/miR-30a-5p/ROR1 axis in modulating chemosensitivity of colorectal cancer cells. We totally collected 294 pairs of colorectal cancer tissues and adjacent normal tissues and also purchased colorectal cancer cell lines and human embryonic kidney cell line. 5-fluorouracil, cisplatin, mitomycin and adriamycin were designated as the chemotherapies for colorectal cell lines, and atractylenolides were arranged as the Chinese drug. The expressions of XIST, miR-30a-5p and ROR1 were quantified with aid of qRT-PCR or Western blot, and luciferase reporter gene assay was implemented to determine the relationships among XIST, miR-30a-5p and ROR1. Our results demonstrated that XIST and ROR1 expressions were dramatically up-regulated, yet miR-30a-5p expression was down-regulated within colorectal cancer tissues (P < 0.05). The overexpressed XIST and ROR1, as well as under-expressed miR-30a-5p, were inclined to promote viability and proliferation of colorectal cells under the influence of chemo drugs (P < 0.05). In addition, XIST could directly target miR-30a-5p, and ROR1 acted as the targeted molecule of miR-30a-5p. Interestingly, atractylenolides not only switched the expressions of XIST, miR-30a-5p and ROR1 within colorectal cancer cells but also significantly intensified the chemosensitivity of colorectal cancer cells (P < 0.05). Finally, atractylenolide II was discovered to slow down the viability and proliferation of colorectal cancer cells (P < 0.05). In conclusion, the XIST/miR-30a-5p/ROR1 axis could be deemed as pivotal markers underlying colorectal cancer, and administration of atractylenolide II might improve the chemotherapeutic efficacy for colorectal cancer.

Amphetamine Neurotoxicity in PC12 Cells through the PP2A/AKT/GSK3ß Pathway.

Amphetamine (AMPH) abuse can influence neuropsychiatric disorders and cell apoptosis by interfering with the protein kinase B/ glycogen synthase kinase 3 beta (AKT/GSK3ß) pathway. However, the mechanisms underlying this regulation are poorly understood. Using PC12 cells, we found that AMPH inhibited AKT and GSK-3ß phosphorylation levels and increased total GSK-3ß levels. Furthermore, AMPH caused an increase in the activity of protein phosphatase 2 (PP2A), a signaling protein upstream of AKT, which in turn inhibited phosphorylated AKT levels. Okadaic acid, a PP2A inhibitor, protected PC12 cells against AMPH-induced apoptosis. Together, our results suggest that the PP2A/AKT/GSK3ß pathway plays an important role in AMPH-induced neurotoxicity.

CSF-Based Analysis for Identification of Potential Serum Biomarkers of Neural Tube Defects.

The protein composition of cerebrospinal fluid (CSF) in neural tube defects (NTDs) remains unknown. We investigated the protein composition of CSF from 9 infants with NTDs using isobaric tags for relative and absolute quantitation (iTRAQ). We identified 568 proteins in the CSF of infants with spina bifida, which is the most common type of NTD. Among these, 18 proteins were associated with neural tube closure in the CSF during human embryonic neurulation and 5 were involved in NTDs. Based on these results, an animal model was further utilized to investigate early serum biomarkers for NTDs. We found that the myristoylated alanine-rich C-kinase substrate, Kunitz-type protease inhibitor 2, and apolipoprotein B-100 protein levels were decreased in both embryos and the sera of pregnant Sprague-Dawley rats carrying embryos with NTDs. CSF proteins may be useful in the discovery of potential serum biomarkers for NTDs.