Emerging Strategies for Modifying Cytochrome P450 Monooxygenases into Peroxizymes.

ConspectusCytochrome P450 monooxygenase is a versatile oxidizing enzyme with great potential in synthetic chemistry and biology. However, the dependence of its catalytic function on the nicotinamide cofactor NAD(P)H and redox partner proteins limits the practical catalytic application of P450 in vitro. An alternative to expensive cofactors is low-cost H2O2, which can be used directly to exploit the catalytic potential of P450s. However, the peroxide shunt pathway is generally inefficient at driving P450 catalysis compared to normal NAD(P)H-dependent activity. Over the last few decades, the scientific community has made continuous efforts to use directed evolution or site-directed mutagenesis to modify P450 monooxygenases into their peroxizyme modes─peroxygenase and peroxidase. Despite significant progress, obtaining efficient P450 peroxizymes remains a huge challenge. Here, we summarize our efforts to modulate peroxizyme activity in P450 monooxygenases and exploit their catalytic applications in challenging selective C-H oxidation, oxygenation, and oxyfunctionalization over the past seven years. We first developed a dual-functional small molecule (DFSM) strategy for transforming P450BM3 monooxygenase into peroxygenase. In this strategy, the typical DFSM, such as N-(ω-imidazolyl)-hexanoyl-l-phenylalanine (Im-C6-Phe), binds to the P450BM3 protein with an anchoring group at one end and plays a general acid-base catalytic role in the activation of H2O2 with an imidazolyl group at the other end. Compared with the O-O homolysis mechanism in the absence of DFSM, the addition of DFSM efficiently enables the heterolytic O-O cleavage of the adduct Fe-O-OH, thus being favored for the formation of active species compound I, which has been demonstrated by combining crystallographic and theoretical calculations. Furthermore, protein engineering showed the unique catalytic performance of DFSM-facilitated P450 peroxygenase for the highly difficult selective oxidation of C-H bonds. This catalytic performance was demonstrated during the chemoselective hydroxylation of gaseous alkanes, regioselective O-demethylation of aryl ethers, highly (R)-enantioselective epoxidation of styrene, and regio- and enantiomerically diverse hydroxylation of alkylbenzenes. Second, we demonstrated that DFSM-facilitated P450BM3 peroxygenase could be effectively switched to an efficient peroxidase mode through mechanism-guided protein engineering of redox-sensitive residues. Utilizing the peroxidase function of P450 enabled the direct nitration of unsaturated hydrocarbons including phenols, aromatic amines, and styrene derivatives, which was not only the P450-catalyzed direct nitration of phenols and aromatic amines for the first time but also the first example of the direct biological nitration of olefins. Finally, we report an H2O2 tunnel engineering strategy to enable peroxygenase activity in several different P450 monooxygenases for the first time, providing a general approach for accessing engineered P450 peroxygenases. In this Account, we highlight the emerging strategies we have developed for producing practical P450 peroxizyme biocatalysts. Although the DFSM strategy is primarily applied to P450BM3 to date, both strategies of redox-sensitive residue engineering and H2O2 tunnel engineering show great potential to extend to other P450s. These strategies have expanded the scope of applications of P450 chemistry and catalysis. Additionally, they provide a unique solution to the challenging selective oxidation of inert C-H bonds in synthetic chemistry.

ABBV-744 induces autophagy in gastric cancer cells by regulating PI3K/AKT/mTOR/p70S6k and MAPK signaling pathways.

The mortality rates of gastric cancer remain high due to limited therapeutic strategies. As a highly selective inhibitor of the BD2 domain of BET family proteins, ABBV-744 has potent chemotherapeutic activity against various human solid tumors. However, whether ABBV-744 has potential anti-tumor effects in gastric cancer remain largely unknown. In this study, we evaluated the effect of ABBV-744 on gastric cancer cells and explored the possible underlying mechanisms. We found that ABBV-744 inhibited the growth of gastric cancer cells and patient-derived tumor organoids in a dose-dependent manner. Cellular experiments revealed that ABBV-744 induced mitochondria damage, reactive oxygen species accumulation, cell cycle arrest and apoptotic cell death in gastric cancer cells. Transcriptomic analysis using RNA-sequencing data identified autophagy as a crucial pathway involved in the cell death caused by ABBV-744. Mechanically, further studies showed that ABBV-744 induced autophagy flux in gastric cancer cells by inactivating PI3K/AKT/mTOR/p70S6k and activating the MAPK signaling pathways. In vivo mouse xenograft studies demonstrated that ABBV-744 significantly suppressed the growth of gastric cancer cells via inducing autophagy. Taken together, our results suggest that ABBV-744 is a novel drug candidate for gastric cancer.

A Unique P450 Peroxygenase System Facilitated by a Dual-Functional Small Molecule: Concept, Application, and Perspective.

Cytochrome P450 monooxygenases (P450s) are promising versatile oxidative biocatalysts. However, the practical use of P450s in vitro is limited by their dependence on the co-enzyme NAD(P)H and the complex electron transport system. Using H2O2 simplifies the catalytic cycle of P450s; however, most P450s are inactive in the presence of H2O2. By mimicking the molecular structure and catalytic mechanism of natural peroxygenases and peroxidases, an artificial P450 peroxygenase system has been designed with the assistance of a dual-functional small molecule (DFSM). DFSMs, such as N-(ω-imidazolyl fatty acyl)-l-amino acids, use an acyl amino acid as an anchoring group to bind the enzyme, and the imidazolyl group at the other end functions as a general acid-base catalyst in the activation of H2O2. In combination with protein engineering, the DFSM-facilitated P450 peroxygenase system has been used in various oxidation reactions of non-native substrates, such as alkene epoxidation, thioanisole sulfoxidation, and alkanes and aromatic hydroxylation, which showed unique activities and selectivity. Moreover, the DFSM-facilitated P450 peroxygenase system can switch to the peroxidase mode by mechanism-guided protein engineering. In this short review, the design, mechanism, evolution, application, and perspective of these novel non-natural P450 peroxygenases for the oxidation of non-native substrates are discussed.

The construction and immunogenicity analyses of a recombinant pseudorabies virus with porcine circovirus type 3 capsid protein co-expression.

Porcine circovirus-associated diseases (PCVADs) and pseudorabies (PR) are highly contagious and economically significant diseases of swine in China. Porcine circovirus type 3 (PCV3) is an emerging swine pathogen of PCVAD. Currently, no PCV3 vaccine is commercially available, and the epidemic caused by it is still spreading worldwide. In this study, we used the PRV variant strain HNX as the parental virus to construct recombinant PRV with TK/gE gene deletion and capsid (Cap) protein co-expression, named HNX-ΔTK/ΔgE-ORF2. The results revealed that PCV3 Cap protein can be detected in HNX-ΔTK/ΔgE-ORF2-infected PK-15 cells by both western blotting and immunofluorescence assays. Vaccination with HNX-ΔTK/ΔgE-ORF2 did not cause pruritus, ruffled fur, systemic infection, or inflammation (without high expression of interleukin-6 (IL-6) and granulocyte colony-stimulating factor (G-CSF) in plasma). Furthermore, HNX-ΔTK/ΔgE-ORF2 immunization induced an anti-Cap specific antibody, activated a PRV-specific cellular immune response, and provided 100 % protection to mice against the challenge of the virulent HNX strain. Thus, HNX-ΔTK/ΔgE-ORF2 appears to be a promising vaccine candidate against PRV and PCV3 for the control of the PRV variant and PCV3.

Akkermansia muciniphila and its outer protein Amuc_1100 regulates tryptophan metabolism in colitis.

Dietary interventions, including dietary ingredients, nutrients and probiotics, exert anti-inflammatory effects in ulcerative colitis (UC). Our previous study showed that Akkermansia muciniphila (Akk), a promising probiotic, could protect against colitis via the regulation of the immune response. However, whether it can restore aberrant tryptophan (Trp) metabolism during colitis remains unclear. In this study, untargeted serum metabolomics of patients with UC and colitis mice showed that Trp metabolism was activated, which was confirmed by quantification of Trp metabolites from a validation cohort and animal study. Integrative analysis of faecal metagenomes and serum metabolomes revealed significant associations between Akk and three Trp metabolites. Live Akk, pasteurised Akk and Amuc_1100 failed to restore the reduction in Trp metabolites involved in the serotonin pathway in colitis mice. However, live Akk, pasteurised Akk and Amuc_1100 increased kynurenine (Kyn) but decreased 2-picolinic acid (PIC) levels and the PIC/Kyn ratio without regulating any of the genes involved in Trp metabolism, suggesting that they could suppress the Kyn pathway (KP) independent of colon tissue. In addition, they could significantly restore the enrichment of Trp metabolism mediated by faecal microbiota. Specifically, live Akk, pasteurised Akk and Amuc_1100 could significantly offset the reduction in indoleacetic acid (IAA) levels. Pasteurised Akk significantly elevated the serum levels of indole acrylic acid (IA). In addition, live Akk, pasteurised Akk and Amuc_1100 could upregulate aryl hydrocarbon receptor (AhR) targeted genes, including CYP1A1, IL-10 and IL-22, suggesting that Akk could activate AhR signaling by regulating Trp metabolism, thereby attenuating colonic inflammation.

Therapeutic potential of an intestinotrophic hormone, glucagon-like peptide 2, for treatment of type 2 short bowel syndrome rats with intestinal bacterial and fungal dysbiosis.

BACKGROUND: Previous studies showed that type 2 short bowel syndrome (SBS) rats were accompanied by severe intestinal bacterial dysbiosis. Limited data are available for intestinal fungal dysbiosis. Moreover, no effective therapeutic drugs are available for these microbiota dysbiosis. The aims of our study were to investigate the therapeutic potential of glucagon-like peptide 2 (GLP-2) for these microbiota dysbiosis in type 2 SBS rats. METHODS: 8-week-old male SD rats which underwent 80% small bowel resection, ileocecum resection, partial colon resection and jejunocolostomy, were treated with saline (SBS group, n = 5) or GLP-2 (GLP2.SBS group, n = 5). The Sham group rats which underwent transection and re-anastomosis were given a saline placebo (Sham group, n = 5). 16S rRNA and ITS sequencing were applied to evaluate the colonic bacterial and fungal composition at 22 days after surgery, respectively. RESULTS: The relative abundance of Actinobacteria, Firmicutes and proinflammatory Proteobacteria increased significantly in SBS group rats, while the relative abundance of Bacteroidetes, Verrucomicrobia and Tenericutes decreased remarkably. GLP-2 treatment significantly decreased Proteus and increased Clostridium relative to the saline treated SBS rats. The diversity of intestinal fungi was significantly increased in SBS rats, accompanied with some fungi abnormally increased and some resident fungi (e.g., Penicillium) significantly decreased. GLP-2 treatment significantly decreased Debaryomyces and Meyerozyma, and increased Penicillium. Moreover, GLP-2 partially restored the bacteria-fungi interkingdom interaction network of SBS rats. CONCLUSION: Our study confirms the bacterial and fungal dysbiosis in type 2 SBS rats, and GLP-2 partially ameliorated these microbiota dysbiosis.

Recombinant Pseudorabies Virus with TK/gE Gene Deletion and Flt3L Co-Expression Enhances the Innate and Adaptive Immune Response via Activating Dendritic Cells.

Owing to viral evolution and recombination, emerging pseudorabies virus (PRV) strains have caused unprecedented outbreaks in swine farms even when the pigs were previously vaccinated, which might indicate that traditional vaccines were unable to provide effective protection. The development of safe and efficacious vaccines presents prospects to minimize the clinical signs and eventually eradicate the infection. In this study, we used an emerging PRV strain, HNX, as the parental strain to construct a recombinant PRV with TK/gE gene deletion and Fms-related tyrosine kinase 3 ligand (Flt3L) expression, named HNX-TK-/gE--Flt3L. HNX-TK-/gE--Flt3L enhanced the maturation of bone marrow derived dendritic cells (DCs) in vitro. Significantly more activated DCs were detected in HNX-TK-/gE--Flt3L-immunized mice compared with those immunized with HNX-TK-/gE-. Subsequently, a remarkable increase of neutralizing antibodies, gB-specific IgG antibodies, and interferon-gamma (IFN-γ) was observed in mice vaccinated with HNX-TK-/gE--Flt3L. In addition, a lower mortality and less histopathological damage were observed in HNX-TK-/gE--Flt3L vaccinated mice with upon PRV lethal challenge infection. Taken together, our results revealed the potential of Flt3L as an ideal adjuvant that can activate DCs and enhance protective immune responses and support the further evaluation of HNX-TK-/gE--Flt3L as a promising PRV vaccine candidate.

Isolation and genetic characteristics of a neurotropic teschovirus variant belonging to genotype 1 in northeast China.

Porcine teschovirus (PTV) is a causative agent of reproductive disorders, encephalomyelitis, respiratory diseases, and diarrhea in swine, with a worldwide distribution. In this work, we identified PTV-associated nonsuppurative encephalitis as a potential cause of posterior paralysis in neonatal pigs in northeast China. Using indirect immunofluorescence assay, western blot, electron microscopy, and genome sequencing, we identified a neurotropic PTV strain, named CHN-NP1-2016, in the supernatants of pooled cerebrum and cerebellum samples from an affected piglet. Nucleotide sequence alignment revealed that the whole genome of CHN-NP1-2016 shared the highest sequence similarity (86.76% identity) with PTV 1 strain Talfan. A combination of phylogenetic and genetic divergence analysis was applied based on the deduced amino acid sequence of the P1 gene with a cutoff value of the genetic distance (0.102 ± 0.008) for defining PTV genotypes, and this showed that CHN-NP1-2016 is a variant of genotype 1. In total, 16 unique mutations and five mutant clusters were detected in the capsid proteins VP1 and VP2 of CHN-NP1-2016 when compared to other PTV1 isolates. Importantly, we detected three mutant clusters located in the exposed surface loops of the capsid protein, potentially indicating significant differences in major neutralization epitopes. Moreover, a potential recombination event in the P1 region of PTV CHN-NP1-2016 was detected. These findings provide valuable insights into the role of recombination in the evolution of teschoviruses. To our knowledge, this is the first case report of PTV-1-associated encephalitis in northeast China. Future investigations will narrow on the serology and pathogenicity of this novel isolate.

Differential expression and correlation analysis of miRNA-mRNA profiles in swine testicular cells infected with porcine epidemic diarrhea virus.

The variant virulent porcine epidemic diarrhea virus (PEDV) strain (YN15) can cause severe porcine epidemic diarrhea (PED); however, the attenuated vaccine-like PEDV strain (YN144) can induce immunity in piglets. To investigate the differences in pathogenesis and epigenetic mechanisms between the two strains, differential expression and correlation analyses of the microRNA (miRNA) and mRNA in swine testicular (ST) cells infected with YN15, YN144, and mock were performed on three comparison groups (YN15 vs Control, YN144 vs Control, and YN15 vs YN144). The mRNA and miRNA expression profiles were obtained using next-generation sequencing (NGS), and the differentially expressed (DE) (p-value < 0.05) mRNA and miRNA were obtained using DESeq R package. mRNAs targeted by DE miRNAs were predicted using the miRanda algortithm. 8039, 8631 and 3310 DE mRNAs, and 36, 36, and 22 DE miRNAs were identified in the three comparison groups, respectively. 14,140, 15,367 and 3771 DE miRNA-mRNA (targeted by DE miRNAs) interaction pairs with negatively correlated expression patterns were identified, and interaction networks were constructed using Cytoscape. Six DE miRNAs and six DE mRNAs were randomly selected to verify the sequencing data by real-time relative quantitative reverse transcription polymerase chain reaction (qRT-PCR). Based on bioinformatics analysis, we discovered the differences were mostly involved in host immune responses and viral pathogenicity, including NF-κB signaling pathway and bacterial invasion of epithelial cells, etc. This is the first comprehensive comparison of DE miRNA-mRNA pairs in YN15 and YN144 infection in vitro, which could provide novel strategies for the prevention and control of PED.

The epidemiological investigation of co-infection of major respiratory bacteria with pseudorabies virus in intensive pig farms in China.

Porcine respiratory disease complex (PRDC), a respiratory disease caused by a variety of factors, is one of the most common problems in the intensive pig farms. To investigate the mixed infection incidence of wild-type pseudorabies virus (WT PRV) and respiratory bacteria, a total of 1,293 clinical samples were collected from pigs with typical respiratory signs from 14 different provinces of China from September 2016 to February 2018. The WT PRV was detected by ELISA targeting gE antibody while the bacteria were detected by bacterial isolation and serotyping by PCR. The results revealed that the detection rate of A. pleuropneumoniae and B. bronchiseptica infection associated with WT PRV infection were 6.30% and 15.99%, respectively, which were significantly higher than those without WT PRV infection (3.41% and 4.41%) at the farm level (p < .05). There were no significant differences in the detection rate of H. parasuis, S. suis or P. multocida between WT PRV positive and negative farms (p > .05). However, the detection rate of attenuated H. parasuis and S. suis strains were 68.19% and 64.75%, respectively, in WT PRV infected farms, which were significantly higher than those (41.56% and 52.25%) in WT PRV free farms (p < .05). The prevalent serotypes of H. parasuis-5/12 and S. suis-2 were also investigated by multiplex PCR. These results indicated that the presence of WT PRV increased the chance of bacterial infection and the number of pathogenic strains in the respiratory system of pigs. Therefore, the eradication of pseudorabies is an effective approach to prevent and control the bacterial respiratory diseases in the intensive pig farms in China.

Establishment of a Blocking ELISA Detection Method for Against African Swine Fever Virus p30 Antibody.

African swine fever (ASF) is a highly lethal hemorrhagic viral disease of domestic pigs caused by African swine fever virus (ASFV). A sensitive and reliable serological diagnostic assay is required, so laboratories can effectively and quickly detect ASFV infection. The p30 protein is abundantly expressed early in cells and has excellent antigenicity. Therefore, this study aimed to produce and characterize p30 monoclonal antibodies with an ultimate goal of developing a monoclonal antibody-based enzyme-linked immunosorbent assay (ELISA) for ASFV antibody detection. Three monoclonal antibodies against p30 protein that were expressed in E. coli were generated, and their characterizations were investigated. Furthermore, a blocking ELISA based on a monoclonal antibody was developed. To evaluate the performance of the assay, 186 sera samples (88 negative and 98 positive samples) were analyzed and a receiver-operating characteristic (ROC) analysis was applied to determine the cutoff value. Based on the ROC analysis, the area under the curve (AUC) was 0.997 (95% confidence interval: 99.2 to 100%). Besides, a diagnostic sensitivity of 97.96% (95% confidence interval: 92.82 to 99.75%) and a specificity of 98.96% (95% confidence interval: 93.83 to 99.97%) were achieved when the cutoff value was set to 38.38%. Moreover, the coefficients of inter- and intra-batches were <10%, indicating the good repeatability of the method. The maximum dilution of positive standard serum detected by this ELISA method was 1:512. The blocking ELISA was able to detect seroconversion in two out of five pigs at 10 Dpi and the p30 response increasing trend through the time course of the study (0-20 Dpi). In conclusion, the p30 mAb-based blocking ELISA developed in this study demonstrated a high repeatability with maximized diagnostic sensitivity and specificity. The assay could be a useful tool for field surveillance and epidemiological studies in swine herd.

PCV2 Regulates Cellular Inflammatory Responses through Dysregulating Cellular miRNA-mRNA Networks.

Porcine circovirus type 2 (PCV2) is closely linked to postweaning multisystemic wasting syndrome (PMWS) and other PCV-associated diseases (PCVADs), which influence the global pig industry. MicroRNAs (miRNAs) are evolutionarily conserved classes of endogenous small non-coding RNA that regulate almost every cellular process. According to our previous transcription study, PCV2 infection causes up-regulation of genes related to inflammation. To reveal the function of miRNAs in PCV2 infection and PCV2-encoded miRNAs, next generation sequencing and data analysis was performed to explore miRNA expression in PCV2-infected cells and non-infected cells. Data analysis found some small RNAs matched the PCV2 genome but PCV2 does not express miRNAs in an in vitro infection (PK-15 cells). More than 297 known and 427 novel miRNAs were identified, of which 44 miRNAs were differently expressed (DE). The pathways of inflammation mediated by chemokine and cytokine signaling pathway (P00031), were more perturbed in PCV2-infected cells than in mock controls. DE miRNAs and DE mRNA interaction network clearly revealed that PCV2 regulates the cellular inflammatory response through dysregulating the cellular miRNA-mRNA network. MiRNA overexpression and down-expression results demonstrated that miRNA dysregulation could affect PCV2 infection-induced cellular inflammatory responses. Our study revealed that host miRNA can be dysregulated by PCV2 infection and play an important role in PCV2-modulated inflammation.

Establishment and application of a multiplex RT-PCR to differentiate wild-type and vaccine strains of porcine epidemic diarrhea virus.

Due to outbreaks of porcine epidemic diarrhea (PED) and the wide use of attenuated live vaccine, both wild-type and vaccine strains (CV777) are believed to circulate in Chinese pig farms. Thus, identification of different PEDV strains is of epidemiological importance. In this study, a multiplex RT-PCR method was established based on the sequence features of spike (S) gene and ORF3 gene of PEDVs. The method could identify PEDV variant strains, classical wild-type strains and classical vaccine strains. The limit of detection of the RT-PCR was 1.51 × 104 copies/uL for plasmids and 1 × 101.7 TCID50/100 u L for PEDV, respectively. There were no cross-detections among three different PEDVs and no false detections among six swine pathogens. This assay was used to test 940 samples from China of which 303 samples were PEDV positive, and 289, 5, 10 were positive for variant, classical wild, classical vaccine, respectively. One sample was positive for both variant and classical vaccine PEDV. The variant PEDVs could be detected in samples from 13 provinces, while classical PEDVs were detected from nine provinces, supporting the prevalence of variant PEDV in China. In summary, this multiplex RT-PCR was a useful tool for the clinical detection and epidemiological survey of PEDV.

Extensive small bowel metastasis and peritoneal dissemination 1 year following curative gastrectomy for T1N1 gastric cancer: A case report.

RATIONALE: Metastasis of T1N1 gastric cancer (GC) at early stage after curative gastrectomy is unusual. Reports on the diagnosis, treatment, and prognosis of peritoneal metastasis following curative gastrectomy for T1N1 GC are lacking. PATIENT CONCERNS: A 54-year-old woman was admitted to our hospital with complaints of mild abdominal distension and failure to pass gas and stool for 2 days. She has a history of distal gastrectomy for T1N1 GC. About 1 year after surgery, she presented with persistent abdominal distension and underwent conservative managements. DIAGNOSES: Imaging tests failed to identify the apparent cause of intestinal obstruction. When conservative managements failed to relieve the symptoms, she underwent emergency laparotomy, which revealed extensive small bowel metastasis and peritoneal dissemination. INTERVENTIONS: Peritoneal irrigation and drainage were performed with the consent of the patient's families. OUTCOMES: The patient abandoned further therapy and died 1 week later during the follow-up period. LESSONS: Although the metastasis of T1N1 GC is rare, patients with high risk of metastasis after curative surgery should also be closely followed and be considered as candidates for more aggressive screening strategies. In addition, the use of more effective chemotherapeutic drugs as adjuvant chemotherapy after curative surgery in T1N1 patients may also need to be explored.

[Establishment and application of gastric and gastric cancer organoids].

Gastric organoid is the organotypic cultures of gastric stem cells or pluripotent stem cells. Gastric organoid is comprised of all major types of gastric epithelial cells and represent the architecture and function remarkably similar to those of the gastric epithelium, faithfully recapitulating the functional gastric epithelium ex vivo. As ideal basic experimental model, gastric organoid has advantages over animal models and conventional cell model in many aspects. Gastric organoid derived from human gastric tissue, in particular, allows the investigation of the function of human stomach in the ex vivo setting. It has now been applied in the field of formation and physiology of the stomach, Helicobacter pylori infection-associated diseases, research of the pathogenic gene, screening and development of drugs, and regenerative medicine. What is more, as an innovative pre-clinical cancer model, gastric cancer organoid has provided important insights in the development of gastric cancer and screening of antitumor drugs, such as simulating the occurrence and development of gastric cancer, screening and development of antitumor drugs, personalized medication and targeted therapy for gastric cancer, and combined application with patient-derived xenograft. In this review, we summarize the establishment and application of gastric and gastric cancer organoids, especially in modeling gastric cancer, basic research and drug development.

Complete Genome Sequence of a Novel Porcine Circovirus Type 3 Strain, PCV3/CN/Hubei-618/2016, Isolated from China.

In October 2016, porcine circovirus type 3 (PCV3) was identified as a pathogen agent for pigs in the United States. Here, we report the genome sequence of a Chinese PCV3 strain, PCV3/CN/Hubei-618/2016. This will help us better understand the epidemiology and genetic characteristics of PCV3.

Porcine Epidemic Diarrhea Virus Induces Autophagy to Benefit Its Replication.

The new porcine epidemic diarrhea (PED) has caused devastating economic losses to the swine industry worldwide. Despite extensive research on the relationship between autophagy and virus infection, the concrete role of autophagy in porcine epidemic diarrhea virus (PEDV) infection has not been reported. In this study, autophagy was demonstrated to be triggered by the effective replication of PEDV through transmission electron microscopy, confocal microscopy, and Western blot analysis. Moreover, autophagy was confirmed to benefit PEDV replication by using autophagy regulators and RNA interference. Furthermore, autophagy might be associated with the expression of inflammatory cytokines and have a positive feedback loop with the NF-κB signaling pathway during PEDV infection. This work is the first attempt to explore the complex interplay between autophagy and PEDV infection. Our findings might accelerate our understanding of the pathogenesis of PEDV infection and provide new insights into the development of effective therapeutic strategies.

High Prevalence of Suboptimal Vitamin D Status and Bone Loss in Adult Short Bowel Syndrome Even After Weaning Off Parenteral Nutrition.

BACKGROUND: Previous studies have noticed the high incidence of suboptimal vitamin D (VtD) status and bone loss in short bowel syndrome (SBS) with parenteral nutrition (PN) dependence. However, limited data have focused on adult SBS without PN dependence. Therefore, our objective was to investigate the incidence and risk factors of suboptimal VtD status and bone loss in adult SBS even after weaning off PN. MATERIALS AND METHODS: We performed a prospective study of 60 adult patients with SBS. Serum 25-hydroxyvitamin D (25-OHD) was measured by radioimmunoassay. Bone mineral density (BMD) was measured using dual-energy x-ray absorptiometry (DEXA). Medical records and various laboratory parameters were collected in all participants. RESULTS: Suboptimal VtD status was identified in all individuals, including 3 (5.0%) with VtD insufficiency and 57 (95.0%) with VtD deficiency. Residual small bowel length (B, 0.072, P = .001) and duration of SBS (B, -0.066, P = .020) were both significantly correlated with suboptimal VtD levels. Overall, only 2 patients presented a normal BMD; osteopenia and osteoporosis were noted in 41 (68.3%) and 17 (28.3%) individuals, respectively. Low 25-OHD concentration was associated with a decreased BMD (B, 0.065, P = .029). There were no other demographic characteristics or clinical examinations associated with suboptimal VtD levels and bone loss. CONCLUSION: Suboptimal VtD status and bone loss were common in adult SBS even after weaning off PN. Despite routine oral VtD supplementation, most patients did not achieve satisfactory status. This emphasizes the critical importance of routine surveillance of 25-OHD and BMD, as well as consideration of alternative methods of supplementation after weaning off PN.

Coordinated Hospital-Home Fecal Microbiota Transplantation via Percutaneous Endoscopic Cecostomy for Recurrent Steroid-Dependent Ulcerative Colitis.

Since its introduction as an alternative intestinal microbiota alteration approach, fecal microbiota transplantation (FMT) has been increasingly used as a treatment of choice for patients with ulcerative colitis (UC), but no reports exist regarding FMT via percutaneous endoscopic cecostomy (PEC). This report describes the case of a 24-year-old man with a 7-year history of recurrent, steroid-dependent UC. He received FMT via PEC once per day for 1 month in the hospital. After the remission of gastrointestinal symptoms, he was discharged from the hospital and continued FMT via PEC twice per week for 3 months at home. The frequency of stools decreased, and the characteristics of stools improved soon thereafter. Enteral nutrition was regained after 1 week, and an oral diet was begun 1 month later. Two months after the FMT end point, the patient resumed a normal diet, with formed soft stools once per day. The follow-up colonoscopy showed normal mucus membranes; then, the PEC set was removed. On the subsequent 12 months follow-up, the patient resumed orthobiosis without any gastrointestinal discomfort and returned to work. This case emphasizes that FMT via PEC can not only induce remission but also shorten the duration of hospitalization and reduce the medical costs; therefore, this approach should be considered an alternative option for patients with UC.

Recurrent scrotal edema in a patient with radiation enteritis: A case report.

Since its introduction as an alternative treatment technique, radiotherapy has been increasingly used as the medical treatment of choice for patients with malignant tumors. However, radiotherapy is associated with a number of common, well-described side effects, which may compromise the quality of life of the patients. Scrotal edema is an infrequent complication in patients who undergo pelvic irradiation, which is suspected to be due to lymphatic obstruction. An extensive literature search found no previous case report describing this complication in patients receiving pelvic radiotherapy. Herein, we present a case of recurrent scrotal edema in a 59-year-old man with prostate cancer and radiation enteritis. Conservative therapy was applied and was successful in relieving the symptoms. To the best of our knowledge, this is the first case report of scrotal edema in a patient with radiation enteritis.