Bacteroides methylmalonyl-CoA mutase produces propionate that promotes intestinal goblet cell differentiation and homeostasis.

Propionate is a short-chain fatty acid that is generated upon microbiome-mediated fiber fermentation in the intestine. By modulating immune and metabolic pathways, propionate exerts many health benefits. Key bacterial species, such as Bacteroides thetaiotaomicron, generate propionate, but the biochemical pathways and specific functions remain undetermined. We identified a gene operon-encoding methylmalonyl-CoA mutase (MCM) that contributes to propionate biosynthesis in B. thetaiotaomicron. Colonization of germ-free mice with wild-type or MCM-deficient strains as well as in vitro examination demonstrated that MCM-mediated propionate production promotes goblet cell differentiation and mucus-related gene expression. Intestinal organoids lacking the propionate receptor, GPR41, showed reduced goblet cell differentiation upon MCM-mediated propionate production. Furthermore, although wild-type B. thetaiotaomicron alleviated DSS-induced intestinal inflammation, this effect was abolished in mice receiving the MCM-deficient strain but restored upon propionate supplementation. These data emphasize the critical role of MCM-mediated propionate biosynthesis in goblet cell differentiation, offering potential pathways to ameliorate colitis.

Epithelial presenilin-1 drives colorectal tumour growth by controlling EGFR-COX2 signalling.

OBJECTIVE: Psen1 was previously characterised as a crucial factor in the pathogenesis of neurodegeneration in patients with Alzheimer's disease. Little, if any, is known about its function in the gut. Here, we uncovered an unexpected functional role of Psen1 in gut epithelial cells during intestinal tumourigenesis. DESIGN: Human colorectal cancer (CRC) and control samples were investigated for PSEN1 and proteins of theγ-secretase complex. Tumour formation was analysed in the AOM-DSS and Apc min/+ mouse models using newly generated epithelial-specific Psen1 deficient mice. Psen1 deficient human CRC cells were studied in a xenograft tumour model. Tumour-derived organoids were analysed for growth and RNA-Seq was performed to identify Psen1-regulated pathways. Tumouroids were generated to study EGFR activation and evaluation of the influence of prostanoids. RESULTS: PSEN1 is expressed in the intestinal epithelium and its level is increased in human CRC. Psen1-deficient mice developed only small tumours and human cancer cell lines deficient in Psen1 had a reduced tumourigenicity. Tumouroids derived from Psen1-deficient Apc min/+ mice exhibited stunted growth and reduced cell proliferation. On a molecular level, PSEN1 potentiated tumour cell proliferation via enhanced EGFR signalling and COX-2 production. Exogenous administration of PGE2 reversed the slow growth of PSEN1 deficient tumour cells via PGE2 receptor 4 (EP4) receptor signalling. CONCLUSIONS: Psen1 drives tumour development by increasing EGFR signalling via NOTCH1 processing, and by activating the COX-2-PGE2 pathway. PSEN1 inhibition could be a useful strategy in treatment of CRC.

The Role of Programmed Necrosis in Colorectal Cancer.

For quite a long time, necrosis was considered a chaotic and unorganized form of cell death. However, studies conducted during the past few decades unveiled multiple types of programmed necrosis, such as necroptosis, pyroptosis and ferroptosis. These types of programmed necrosis have been shown to play crucial roles in mediating pathological processes, including tumorigenesis. Almost all key mediators, such as RIPK3 and MLKL in necroptosis, GSDMD and caspase 1/11 in pyroptosis and GPX4 in ferroptosis, are highly expressed in intestinal epithelial cells (IECs). An aberrant increase or decrease in programmed necrosis in IECs has been connected to intestinal disorders. Here, we review the pathways of programmed necrosis and the specific consequences of regulated necrosis in colorectal cancer (CRC) development. Translational aspects of programmed necrosis induction as a novel therapeutic alternative against CRC are also discussed.

SMYD2 Inhibition Downregulates TMPRSS2 and Decreases SARS-CoV-2 Infection in Human Intestinal and Airway Epithelial Cells.

The COVID-19 pandemic caused by SARS-CoV-2 has lasted for more than two years. Despite the presence of very effective vaccines, the number of virus variants that escape neutralizing antibodies is growing. Thus, there is still a need for effective antiviral treatments that target virus replication independently of the circulating variant. Here, we show for the first time that deficiency or pharmacological inhibition of the cellular lysine-methyltransferase SMYD2 decreases TMPRSS2 expression on both mRNA and protein levels. SARS-CoV-2 uses TMPRSS2 for priming its spike protein to infect target cells. Treatment of cultured cells with the SMYD2 inhibitors AZ505 or BAY598 significantly inhibited viral replication. In contrast, treatment of Vero E6 cells, which do not express detectable amounts of TMPRSS2, had no effect on SARS-CoV-2 infection. Moreover, by generating a recombinant reporter virus that expresses the spike protein of the Delta variant of SARS-CoV-2, we demonstrate that BAY598 exhibits similar antiviral activity against this variant of concern. In summary, SMYD2 inhibition downregulates TMPRSS2 and blocks viral replication. Targeting cellular SMYD2 represents a promising tool to curtail SARS-CoV-2 infection.

SMYD2 targets RIPK1 and restricts TNF-induced apoptosis and necroptosis to support colon tumor growth.

SMYD2 is a histone methyltransferase, which methylates both histone H3K4 as well as a number of non-histone proteins. Dysregulation of SMYD2 has been associated with several diseases including cancer. In the present study, we investigated whether and how SMYD2 might contribute to colorectal cancer. Increased expression levels of SMYD2 were detected in human and murine colon tumor tissues compared to tumor-free tissues. SMYD2 deficiency in colonic tumor cells strongly decreased tumor growth in two independent experimental cancer models. On a molecular level, SMYD2 deficiency sensitized colonic tumor cells to TNF-induced apoptosis and necroptosis without affecting cell proliferation. Moreover, we found that SMYD2 targeted RIPK1 and inhibited the phosphorylation of RIPK1. Finally, in a translational approach, pharmacological inhibition of SMYD2 attenuated colonic tumor growth. Collectively, our data show that SMYD2 is crucial for colon tumor growth and inhibits TNF-induced apoptosis and necroptosis.

E-type prostanoid receptor 4 drives resolution of intestinal inflammation by blocking epithelial necroptosis.

Inflammatory bowel diseases present with elevated levels of intestinal epithelial cell (IEC) death, which compromises the gut barrier, activating immune cells and triggering more IEC death. The endogenous signals that prevent IEC death and break this vicious cycle, allowing resolution of intestinal inflammation, remain largely unknown. Here we show that prostaglandin E2 signalling via the E-type prostanoid receptor 4 (EP4) on IECs represses epithelial necroptosis and induces resolution of colitis. We found that EP4 expression correlates with an improved IBD outcome and that EP4 activation induces a transcriptional signature consistent with resolution of intestinal inflammation. We further show that dysregulated necroptosis prevents resolution, and EP4 agonism suppresses necroptosis in human and mouse IECs. Mechanistically, EP4 signalling on IECs converges on receptor-interacting protein kinase 1 to suppress tumour necrosis factor-induced activation and membrane translocation of the necroptosis effector mixed-lineage kinase domain-like pseudokinase. In summary, our study indicates that EP4 promotes the resolution of colitis by suppressing IEC necroptosis.

PGAM5-MAVS interaction regulates TBK1/ IRF3 dependent antiviral responses.

Viral infections trigger host innate immune responses, characterized by the production of type-I interferons (IFN) including IFNß. IFNß induces cellular antiviral defense mechanisms and thereby contributes to pathogen clearance. Accumulating evidence suggests that mitochondria constitute a crucial platform for the induction of antiviral immunity. Here we demonstrate that the mitochondrial protein phosphoglycerate mutase family member 5 (PGAM5) is important for the antiviral cellular response. Following challenge of HeLa cells with the dsRNA-analog poly(I:C), PGAM5 oligomers and high levels of PGAM5 were found in mitochondrial aggregates. Using immunoprecipitation, a direct interaction of PGAM5 with the mitochondrial antiviral-signaling protein (MAVS) was demonstrated. In addition, PGAM5 deficient cells showed diminished expression of IFNß and IFNß target genes as compared to WT cells. Moreover, PGAM5 deficient mouse embryonic fibroblasts (MEFs) exhibited decreased phosphorylation levels of IRF3 and TBK1 when challenged with poly(I:C) intracellularly. Finally, PGAM5 deficient MEFs, upon infection with vesicular stomatitis virus (VSV), revealed diminished IFNß expression and increased VSV replication. Collectively, our study highlights PGAM5 as an important regulator for IFNß production mediated via the TBK1/IRF3 signaling pathway in response to viral infection.

Regression of apoptosis-resistant colorectal tumors by induction of necroptosis in mice.

Cancer cells often acquire capabilities to evade cell death induced by current chemotherapeutic treatment approaches. Caspase-8, a central initiator of death receptor-mediated apoptosis, for example, is frequently inactivated in human cancers via multiple mechanisms such as mutation. Here, we show an approach to overcome cell death resistance in caspase-8-deficient colorectal cancer (CRC) by induction of necroptosis. In both a hereditary and a xenograft mouse model of caspase-8-deficient CRC, second mitochondria-derived activator of caspase (SMAC) mimetic treatment induced massive cell death and led to regression of tumors. We further demonstrate that receptor-interacting protein kinase 3 (RIP3), which is highly expressed in mouse models of CRC and in a subset of human CRC cell lines, is the deciding factor of cancer cell susceptibility to SMAC mimetic-induced necroptosis. Thus, our data implicate that it may be worthwhile to selectively evaluate the efficacy of SMAC mimetic treatment in CRC patients with caspase-8 deficiency in clinical trials for the development of more effective personalized therapy.

Bovine TLR2 and TLR4 mediate Cryptosporidium parvum recognition in bovine intestinal epithelial cells.

Cryptosporidium parvum (C. parvum) is an intestinal parasite that causes diarrhea in neonatal calves. It results in significant morbidity of neonatal calves and economic losses for producers worldwide. Innate resistance against C. parvum is thought to depend on engagement of pattern recognition receptors. However, the role of innate responses to C. parvum has not been elucidated in bovine. The aim of this study was to evaluate the role of TLRs in host-cell responses during C. parvum infection of cultured bovine intestinal epithelial cells. The expressions of TLRs in bovine intestinal epithelial cells were detected by qRT-PCR. To determine which, if any, TLRs may play a role in the response of bovine intestinal epithelial cells to C. parvum, the cells were stimulated with C. parvum and the expression of TLRs were tested by qRT-PCR. The expression of NF-κB was detected by western blotting. Further analyses were carried out in bovine TLRs transfected HEK293 cells and by TLRs-DN transfected bovine intestinal epithelial cells. The results showed that bovine intestinal epithelial cells expressed all known TLRs. The expression of TLR2 and TLR4 were up-regulated when bovine intestinal epithelial cells were treated with C. parvum. Meanwhile, C. parvum induced IL-8 production in TLR2 or TLR4/MD-2 transfected HEK293 cells. Moreover, C. parvum induced NF-κB activation and cytokine expression in bovine intestinal epithelial cells. The induction of NF-κB activation and cytokine expression by C. parvum were reduced in TLR2-DN and TLR4-DN transfected cells. The results showed that bovine intestinal epithelial cells expressed all known TLRs, and bovine intestinal epithelial cells recognized and responded to C. parvum via TLR2 and TLR4.

Thymol attenuates allergic airway inflammation in ovalbumin (OVA)-induced mouse asthma.

Thymol, a naturally occurring monocyclic phenolic compound derived from Thymus vulgaris (Lamiaceae), has been reported to exhibit anti-inflammatory property in vivo and vitro. However, the mechanism of thymol is not clear. The aim of the present study was to investigate the effects of thymol on allergic inflammation in OVA-induced mice asthma and explore its mechanism. The model of mouse asthma was established by the induction of OVA. Thymol was orally administered at a dose of 4, 8, and 16 mg/kg body weight 1h before OVA challenge. At 24h after the last challenge, mice were sacrificed, and the data were collected by various experimental methods. The results revealed that pretreatment with thymol reduced the level of OVA-specific IgE, inhibited recruitment of inflammatory cells into airway, and decreased the levels of IL-4, IL-5, and IL-13 in BALF. Moreover, the pathologic changes of lung tissues were obviously ameliorated and goblet cell hyperplasia was effectively inhibited by the pretreatment of thymol. In addition, thymol reduced the development of airway hyperresponsiveness and blocked the activation of NF-κB pathway. All data suggested that thymol ameliorated airway inflammation in OVA-induced mouse asthma, possibly through inhibiting NF-κB activation. These findings indicated that thymol may be used as an alternative agent for treating allergic asthma.

Thymol inhibits Staphylococcus aureus internalization into bovine mammary epithelial cells by inhibiting NF-κB activation.

Bovine mastitis is one of the most costly and prevalent diseases in the dairy industry and is characterised by inflammatory and infectious processes. Staphylococcus aureus (S. aureus), a Gram-positive organism, is a frequent cause of subclinical, chronic mastitis. Thymol, a monocyclic monoterpene compound isolated from Thymus vulgaris, has been reported to have antibacterial properties. However, the effect of thymol on S. aureus internalization into bovine mammary epithelial cells (bMEC) has not been investigated. In this study, we evaluated the effect of thymol on S. aureus internalization into bMEC, the expression of tracheal antimicrobial peptide (TAP) and ß-defensin (BNBD5), and the inhibition of NF-κB activation in bMEC infected with S. aureus. Our results showed that thymol (16-64 µg/ml) could reduce the internalization of S. aureus into bMEC and down-regulate the mRNA expression of TAP and BNBD5 in bMEC infected with S. aureus. In addition, thymol was found to inhibit S. aureus-induced nitric oxide (NO) production in bMEC and suppress S. aureus-induced NF-κB activation in a dose-dependent manner. In conclusion, these results indicated that thymol inhibits S. aureus internalization into bMEC by inhibiting NF-κB activation.

Molecular characterization of Giardia duodenalis isolates from police and farm dogs in China.

To assess the potential zoonotic transmission of giardiasis from dogs in China, a total of 205 fecal specimens from dogs were screened for Giardia duodenalis using PCR and sequence analysis of the triosephosphate isomerase gene. The prevalence of G. duodenalis in dogs was 13.2% (27/205). The potentially zoonotic assemblage A and the dog-specific assemblage C was identified in 25 (12.2%) and two (1.0%) dogs, respectively. All assemblage A isolates belonged to sub-assemblage AI, genotype AI-1. Likewise, one subtype was found in assemblage C. The high occurrence of potentially zoonotic G. duodenalis subtype AI-1 in dogs that are in close contact with humans is of public health concern.