Robust three-dimensional expansion of human adult alveolar stem cells and SARS-CoV-2 infection

Severe acute respiratory syndrome-coronavirus 2 (SARS-CoV-2), which is the cause of a present global pandemic, infects human lung alveolar cells (hACs). Characterising the pathogenesis is crucial for developing vaccines and therapeutics. However, the lack of models mirroring the cellular physiology and pathology of hACs limits the study. Here, we develop a feeder-free, long-term three-dimensional (3D) culture technique for human alveolar type 2 (hAT2) cells, and investigate infection response to SARS-CoV-2. By imaging-based analysis and single-cell transcriptome profiling, we reveal rapid viral replication and the increased expression of interferon-associated genes and pro-inflammatory genes in infected hAT2 cells, indicating robust endogenous innate immune response. Further tracing of viral mutations acquired during transmission identifies full infection of individual cells effectively from a single viral entry. Our study provides deep insights into the pathogenesis of SARS-CoV-2, and the application of long-term 3D hAT2 cultures as models for respiratory diseases.

Organoids as a New In Vitro Model of Human Norovirus Infection

Human noroviruses (HuNoVs) are the main causative viral agents in epidemic and endemic acute gastroenteritis worldwide. Despite their impact on the global economic and health burden, developing effective control measures to prevent and treat norovirus gastroenteritis remains a difficult problem to solve. One of the major reasons for this problem is the lack of affordable small animal models and a robust and reproducible In Vitro cell culture system for the propagation of this poorly characterized RNA virus. Recently, the development of a 3-dimensional culture system using pluripotent stem cells to mimic the native small intestine has led to the discovery of new strategies for the cultivation of this virus. In this review, we describe a human stem cell-derived intestinal organoid model that led to the development of the currently available HuNoV In Vitroculture systems that support replication of the virus, and provide helpful insights into HuNoV biology and vaccine and therapeutic development.

Oxidative stress and glutamate excretion in alcoholic steatosis: Metabolic synapse between hepatocyte and stellate cell

Chronic alcohol consumption induces the development of alcoholic steatosis in the liver, which is one of the most widespread liver diseases worldwide. During general alcohol metabolism, hepatocytes generate mitochondria- and cytochrome P450 2E1 (CYP2E1)-mediated reactive oxygen species (ROS) whose accumulation elicits activation of the hepatic anti-oxidant system, including glutathione (GSH). However, chronic alcohol consumption decreases GSH generation through cysteine deficiency by suppressing the methionine cycle and trans-sulfuration system, whereas it turns on an alternative defense pathway, such as the xCT transporter, to compensate for GSH shortage. The xCT transporter mediates the uptake of cystine coupled to the efflux of glutamate, leading to an increase in blood glutamate. In response to the elevated glutamate in the liver, the expression of metabotropic glutamate receptor 5 (mGluR5) is up-regulated in hepatic stellate cells (HSCs) along with enhanced production of 2-arachidonoylglycerol, which in turn stimulates cannabinoid receptor 1 (CB1R) on neighboring hepatocytes to increase de novo lipogenesis. On the other hand, blockade of mGluR5 and CB1R attenuates alcoholic steatosis. Interestingly, although the increased expression of CYP2E1-mediated xCT and ROS generation are mainly observed at the perivenous area (zone 3), fat accumulation is mostly detected at hepatic zone 2. To resolve this discrepancy, this review summarizes recent advances on glutamate/mGluR5-derived alcoholic steatosis and zone-dependently different responses to alcohol intake. In addition, the bidirectional loop pathway and its unique metabolic synapse between hepatocytes and HSCs are discussed.

Characterization of Severe Acute Respiratory Syndrome Coronavirus 2Stability in Multiple Water Matrices

The advent of the global pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) necessitates a thorough study of the stability and transmissibility in the environment. We characterized the stability of SARS-CoV-2 in three water matrices: fresh, tap, and seawater. The minimum infective dose of SARS-CoV-2 in Vero cells was confirmed to be 10 3 PFU/mL. The stability of SARS-CoV-2 varied according to the water matrix: infective SARSCoV-2 was undetectable after treatment with fresh water and seawater, but remained detectable for 2 days in tap water, when starting with an initial concentration of 10 4 PFU/mL. When the starting concentration was increased to 10 5 PFU/mL, a similar trend was observed. In addition, viral RNA persisted longer than infectious virus in all water matrices. This study was conducted in stagnant water containing a significantly high titer of virus, thus, human-to-human transmission of SARS-CoV-2 through the actual aquatic environment is expected to be rare.

Organoids as a New In Vitro Model of Human Norovirus Infection

Human noroviruses (HuNoVs) are the main causative viral agents in epidemic and endemic acute gastroenteritis worldwide. Despite their impact on the global economic and health burden, developing effective control measures to prevent and treat norovirus gastroenteritis remains a difficult problem to solve. One of the major reasons for this problem is the lack of affordable small animal models and a robust and reproducible In Vitro cell culture system for the propagation of this poorly characterized RNA virus. Recently, the development of a 3-dimensional culture system using pluripotent stem cells to mimic the native small intestine has led to the discovery of new strategies for the cultivation of this virus. In this review, we describe a human stem cell-derived intestinal organoid model that led to the development of the currently available HuNoV In Vitroculture systems that support replication of the virus, and provide helpful insights into HuNoV biology and vaccine and therapeutic development.

Distribution of Pathogenic Vibrio Species in the Coastal Seawater of South Korea (2017–2018)

OBJECTIVES: Pathogenic Vibrio species are widely distributed in warm estuarine and coastal environments, and can infect humans through the consumption of raw or mishandled contaminated seafood and seawater. For this reason, the distribution of these bacteria in South Korea was investigated.METHODS: Seawater samples were collected from 145 coastal area points in the aquatic environment in which Vibrio species live. Environmental data (i.e., water temperature, salinity, turbidity, and atmospheric temperature) was collected which may help predict the distribution of the species (data not shown). Seawater samples were filtered, and incubated overnight in alkaline peptone water, at 37°C. Using species-specific polymerase chain reaction methods, screening tests were performed for the hlyA, ctxA, vvhA, and tlh genes. Clones of pathogenic Vibrio species were isolated using 3 selective plating media.RESULTS: In 2017, total seawater isolation rates for Vibrio vulnificus, Vibrio cholerae (non-pathogenic, non-O1, non-O139 serogroups), and Vibrio parahaemolyticus were 15.82%, 13.18%, 65.80%, respectively. However, in 2018 isolation rates for each were 21.81%, 19.40%, and 70.05%, respectively.CONCLUSION: The isolation rates of pathogenic Vibrio species positively correlated with the temperature of seawater and atmosphere, but negatively correlated with salinity and turbidity. From 2017 to 2018, the most frequent seawater-isolated Vibrio species were V. parahaemolyticus (68.10 %), V. vulnificus (16.54%), and non-toxigenic V. cholerae (19.58%). Comprehensive monitoring, prevention, and control efforts are needed to protect the public from pathogenic Vibrio species.

Effect of genetic background and delivery route on the preclinical properties of a live attenuated RSV vaccine.

A safe and effective vaccine against RSV remains an important unmet public health need. Intranasally (IN) delivered live-attenuated vaccines represent the most extensively studied approach for immunization of RSV-naïve infants and children, however, achieving an effective balance of attenuation and immunogenicity has proven challenging. Here we report pre-clinical immunogenicity and efficacy data utilizing a live-attenuated vaccine candidate, RGΔM2-2, which was obtained by deleting the M2-2 open reading frame from the genome of the MSA1 clinical isolate. Intramuscular (IM) administration of RGΔM2-2 in cotton rats induced immunity and protective efficacy that was comparable to that induced by intranasal (IN) immunization. In contrast, the protective efficacy of RGΔM2-2 delivered by the IM route to African green monkeys was substantially reduced as compared to the efficacy following IN administration, despite comparable levels of serum neutralizing antibodies. This result suggests that mucosal immunity may play an important role in RSV protection. The RGΔM2-2 vaccine also demonstrated different attenuation profiles when tested in cotton rats, non-human primates, and a human airway epithelial (HAE) cell model. The data suggest RGΔM2-2 is less attenuated than a similarly designed vaccine candidate constructed on the A2 genetic background. These findings have important implications with regard to both the design and the preclinical safety testing of live-attenuated vaccines.

Anti-hyperglycemic effects and signaling mechanism of Perilla frutescens sprout extract

BACKGROUND/OBJECTIVES: Perilla frutescens (L.) Britton var. (PF) sprout is a plant of the labiate family. We have previously reported the protective effects of PF sprout extract on cytokine-induced β-cell damage. However, the mechanism of action of the PF sprout extract in type 2 diabetes (T2DM) has not been investigated. The present study was designed to study the effects of PF sprout extract and signaling mechanisms in the T2DM mice model using C57BL/KsJ-db/db (db/db) mice. MATERIALS/METHODS: Male db/db mice were orally administered PF sprout extract (100, 300, and 1,000 mg/kg of body weight) or rosiglitazone (RGZ, positive drug, 1 mg/kg of body weight) for 4 weeks. Signaling mechanisms were analyzed using liver tissues and HepG2 cells. RESULTS: The PF sprout extract (300 and 1,000 mg/kg) significantly reduced the fasting blood glucose, serum insulin, triglyceride and total cholesterol levels in db/db mice. PF sprout extract also significantly improved glucose intolerance and insulin sensitivity, decreased hepatic gluconeogenic protein expression, and ameliorated histological alterations of the pancreas and liver. Levels of phosphorylated AMP-activated protein kinase (AMPK) protein expression also increased in the liver after treatment with the extract. In addition, an increase in the phosphorylation of AMPK and decrease in the phosphoenolpyruvate carboxykinase and glucose 6-phosphatase proteins in HepG2 cells were also observed. CONCLUSIONS: Our results sugges that PF sprout displays beneficial effects in the prevention and treatment of type 2 diabetes via modulation of the AMPK pathway and inhibition of gluconeogenesis in the liver.

A Case of Intramural Hematoma of the Esophagus Mimicking Acute Coronary Syndrome / 대한소화기학회지

Intramural hematoma of the esophagus is a rare condition that can be spontaneous or secondary to trauma, toxic ingestion, or intervention. If it is the spontaneous type, it usually presents initially with epigastric pain, hematemesis or dysphagia. We present a case of intramural hematoma of the esophagus mimicking acute coronary syndrome. A 63-year-old man presented with severe acute chest pain. He has four coronary stents that were inserted five years ago, from a different hospital, and is on dual antiplatelet agents. Coronary angiography was performed immediately under the suspicion of acute coronary syndrome, and we found that there was no obvious clogging of the coronary arteries. Next, chest computed tomography was performed due to suspected aortic dissection, and the result was also negative. Four days later, endoscopy was performed and intramural hematoma covered with large ulcers was diagnosed.

Serological and Molecular Detection of Toxoplasma gondii and Babesia microti in the Blood of Rescued Wild Animals in Gangwon-do (Province), Korea

Infections of Toxoplasma gondii and Babesia microti are reported in many wild animals worldwide, but information on their incidence and molecular detection in Korean wild fields is limited. In this study, the prevalence of T. gondii and B. microti infection in blood samples of 5 animal species (37 Chinese water deer, 23 raccoon dogs, 6 roe deer, 1 wild boar, and 3 Eurasian badgers) was examined during 2008–2009 in Gangwon-do (Province), the Republic of Korea (=Korea) by using serological and molecular tests. The overall seropositivity of T. gondii was 8.6% (6/70); 10.8% in Chinese water deer, 4.3% in raccoon dogs, and 16.7% in roe deer. PCR revealed only 1 case of T. gondii infection in Chinese water deer, and phylogenic analysis showed that the positive isolate was practically identical to the highly pathogenetic strain type I. In B. microti PCR, the positive rate was 5.7% (4/70), including 2 Chinese water deer and 2 Eurasian badgers. Phylogenetic analysis results of 18S rRNA and the β-tubulin gene showed that all positive isolates were US-type B. microti. To our knowledge, this is the first report of B. microti detected in Chinese water deer and Eurasian badger from Korea. These results indicate a potentially high prevalence of T. gondii and B. microti in wild animals of Gangwon-do, Korea. Furthermore, Chinese water deer might act as a reservoir for parasite infections of domestic animals.

Prevalence of Clonorchis sinensis Infection among Residents along 5 Major Rivers in the Republic of Korea

Clonorchis sinensis is currently the most important parasite affecting public health problems in the Republic of Korea. We investigated the prevalence of C. sinensis infection among residents living along 5 major rivers in Korea. A total of 42,562 individual stool samples were collected from 37 localities and examined using the formalin-ether sedimentation technique. Helminth eggs were detected in 4,052 (9.5%) residents and 3,586 (8.4%) were infected with C. sinensis. The egg positive rate of C. sinensis in Nakdong, Seomjin, Geum, Yeongsan, and Han River was 11.7%, 9.9%, 6.5%, 3.1%, and 1.0%, respectively. The overall prevalence of clonorchiasis by sex was 11.2% in males and 6.2% in females. The age-prevalence was the highest in the 50-59 years band. It has been reconfirmed that the endemicity of clonorchiasis is higher in southern areas of Korea, especially along Nakdong and Seomjin Rivers. A combination of continuous control programs with health education initiatives is urgently required in these highly endemic areas of clonorchiasis in Korea.

Expression and clinical significance of defensin alpha 6 in colorectal cancer

PURPOSE: Defensin alpha 6 (DEFA6) is cationic short peptide with known functional activities in innate antimicrobial immunity. DEFA6 is also highly expressed in colorectal cancer tissue. The mechanism and function of DEFA6 have not been reported how to play a significant role in carcinogenesis and cancer progression. The aim of this study was to evaluate the protumorigenic functions of DEFA6 in the colorectal cancer cell line and the clinical significance of DEFA6 expression in colorectal cancer patients.METHODS: DEFA6 expression was investigated by immunohistochemistry in 151 cases of colorectal cancer tissue and the association of DEFA6 expression was correlated with patient's cancer charecteristics.RESULTS: Immunohistochemistry analysis showed that the DEFA6 protein was expressed higher in advanced cancer group (high T stage, patients with lymph node metastasis, patients with vascular invasion) than early cancer group (low T stage, patients without lymph node metastasis, patients without vascular invasion) (P=0.007/0.032/0.047).CONCLUSION: As patient's survival usually depends on migration and invasion of the cancer cell, the high expression of DEFA6 in colorectal cancer cell is associated with patient's cancer charecteristics and could be a biomarker for colorectal cancer. The future study about characterization of DEFA6 will clarify its specific role in oncogenesis and its therapeutic potential.

Toxoplasma gondii Infection Suppresses House Dust Mite Extract-Induced Atopic Dermatitis in NC/Nga Mice

PURPOSE: Toxoplasma gondii (T. gondii) is an obligate intracellular protozoan parasite that infects humans and animals via congenital or postnatal routes, and it is found worldwide. Modulation of the immune system by parasite infection is proposed to suppress allergic inflammation. Growing evidences have shown that interleukin (IL)-10-producing regulatory B cells (B(regs)) and CD4+CD25+FoxP3+ regulatory T cells (T(regs)) induced by parasite infection play a critical role in allergic or autoimmune diseases because these cells regulate negatively cellular immune responses and inflammation. Currently, the role of IL-10-producing regulatory B cells in host immune response during T. gondii infection is unknown. In this study, we investigate whether T. gondii infection can suppress the development of unrelated atopic dermatitis (AD)-like lesions. METHODS: AD is a chronically relapsing inflammatory skin disease accompanied by severe itching; for this, we used NC/Nga mice, a well-known experimental model of systemic AD. Repeated exposure to Dermatophagoides farinae crude extract (DfE), known as a major environmental allergen, evokes AD-like skin lesions in NC/Nga mice under specific pathogen-free conditions. NC/Nga mice were intraperitoneally infected with 10 cysts of T. gondii. RESULTS: T. gondii infection significantly ameliorated AD-like skin lesions in NC/Nga mice. The subpopulation of B(regs) and T(regs) in the AD mice was expanded in the course of T. gondii infection. In addition, T. gondii infection inhibited Th2 and enhanced Th1 immune response in the DfE-treated AD mice. CONCLUSIONS: We have experimentally demonstrated for the first time that T. gondii infection ameliorated AD-like skin lesions in a mouse model of AD. Our study could in part explain the mechanisms of how parasite infection prevents the development of allergic disorder. Therefore, these immunemechanisms induced by T. gondii infection may be beneficial for the host in terms of reduced risk of allergic immune reactions.

A Case of Long Term Survival in Patient with Early Intrahepatic Recurrence and Extrahepatic Metastasis after Curative Resection of Hepatocellular Carcinoma

The most effective therapy in patients with hepatocellular carcinoma (HCC) is curative resection of liver cancer. However, the long-term prognosis is not good in high intrahepatic recurrence. The recurrence rate after curative resection of HCC is 70-100% and the average survival time was about 15.8 months in early relapse after curative resection. Extra hepatic metastases in HCC is associated not uncommon, in general, lungs are the most common organ, lymph nodes, bone, and adrenal gland is relatively often observed. Extrahepatic recurrence after curative resection of HCC has been reported in 14-26% of the total recurrent cancer. Most of the recurrent cancer is multiple and the median survival time of patients after the extrahepatic recurrence is about 4-5 months it found that the prognosis is very poor. Therefore, the authors report a case showed long term survival in patients with early intrahepatic recur and extrahepatic metastasis after curative resection of HCC.

Trigonelline protects the cardiocyte from hydrogen peroxide induced apoptosis in H9c2 cells

Objective: To elucidate the key parameters associated with hydrogen peroxide induced oxidative stress and investigates the mechanism of trigonelline (TG) for reducing the H2O2 induced toxicity in H9c2 cells. Methods: Cytotoxicity and antioxidant activity of TG was assessed by EZ-CYTOX kit. RNA extraction and cDNA synthesized according to the kit manufacture protocol. Apoptosis was measured by the Flowcytometry, general PCR and qPCR. Results: It was found that the TG significantly rescued the morphology of the H9c2 cells. Treatment of cells with TG attenuated H2O2 induced cell deaths and improved the antioxidant activity. In addition, TG regulated the apoptotic gene caspase-3, caspase-9 and anti-apoptotic gene Bcl-2, Bcl-XL during H2O2 induced oxidative stress in H9c2 cells. These results were comparable with quercetin treatment. For evident, flow cytometer results also confirmed the TG significantly reduced the H2O2 induced necrosis and apoptosis in H9c2 cells. However, further increment of TG concentration against H2O2 could induce the necrosis and apoptosis along with H2O2. Conclusions: It is suggested that less than 125 μ M of TG could protect the cells from H2O2 induced cell damage by down regulating the caspases and up regulating the Bcl-2 and Bcl-XL expression. Therefore, we suggest the trigonelline could be useful for treatment of oxidative stress mediated cardiovascular diseases in future.

Trigonelline protects the cardiocyte from hydrogen peroxide induced apoptosis in H9c2 cells

OBJECTIVE@#To elucidate the key parameters associated with hydrogen peroxide induced oxidative stress and investigates the mechanism of trigonelline (TG) for reducing the H2O2 induced toxicity in H9c2 cells.@*METHODS@#Cytotoxicity and antioxidant activity of TG was assessed by EZ-CYTOX kit. RNA extraction and cDNA synthesized according to the kit manufacture protocol. Apoptosis was measured by the Flowcytometry, general PCR and qPCR.@*RESULTS@#It was found that the TG significantly rescued the morphology of the H9c2 cells. Treatment of cells with TG attenuated H2O2 induced cell deaths and improved the antioxidant activity. In addition, TG regulated the apoptotic gene caspase-3, caspase-9 and anti-apoptotic gene Bcl-2, Bcl-XL during H2O2 induced oxidative stress in H9c2 cells. These results were comparable with quercetin treatment. For evident, flow cytometer results also confirmed the TG significantly reduced the H2O2 induced necrosis and apoptosis in H9c2 cells. However, further increment of TG concentration against H2O2 could induce the necrosis and apoptosis along with H2O2.@*CONCLUSIONS@#It is suggested that less than 125 μ M of TG could protect the cells from H2O2 induced cell damage by down regulating the caspases and up regulating the Bcl-2 and Bcl-XL expression. Therefore, we suggest the trigonelline could be useful for treatment of oxidative stress mediated cardiovascular diseases in future.

Colonic Diverticulitis Complicating Colo-ovarian Fistula and Tuboovarian Abscess Formation in on Old Woman

The inflammation of colonic diverticulum, diverticulitis, is occasionally accompanied by several complications, such as bleeding, bowel obstruction, fistula and abscess formation. Both colo-ovarian fistula and tuboovarian abscess formation, as complications of diverticulitis, are rare. Here, we report a case of colonic diverticulitis which ultimately resulted in colo-ovarian fistula and tuboovarian abscess formation. A 76-year-old female was presented with lower abdominal pain, diarrhea and intermittent hematochezia for about 1 month. According to the abdominal computed tomography, tuboovarian abscess was shown to adhere to the inflamed sigmoid diverticulum. Sigmoidoscopy revealed pus flowing from the opening of the diverticulum at the sigmoid colon. Hence, the patient underwent oophorosalpingectomy and low anterior resection. Thereafter, she was treated with bowel rest and broad spectrum antibiotics, and finally recovered. Although rare, colonic diverticulitis may result in colo-ovarian fistula causing tuboovarian abscess formation in postmenopausal patients. Early recognition of the condition and appropriate treatment, including surgery and antibiotics, are necessary in order to achieve full recovery from complicated diverticulitis.

Clinical Analysis of Risk Factors for Recurrence in Patients with Chronic Subdural Hematoma Undergoing Burr Hole Trephination / 대한신경손상학회지

OBJECTIVE: Chronic subdural hematoma (CSDH) is one of the most common types of intra-cranial hemorrhages usually associated with trauma. Surgical treatment is the treatment of choice and burr hole trephination (BHT) is widely performed. The recurrence rate in the patients with CSDH is 3.7-30%. This study investigated the risk factors associated with the recurrence of patients with CSDH who underwent BHT. METHODS: One hundred twenty-five patients with CSDH underwent BHT. Eight of 125 patients (6.4%) underwent reoperation for recurrent CSDH. We retrospectively analyzed demographic, clinical and radiological findings, catheter tip location and drainage duration as the risk factors for the recurrence of CSDH. RESULTS: Recurrence of CSDH in the high- or mixed-density groups was significantly higher than those in the low- or iso-density groups (p<0.001). Placement of catheter tip at the temporoparietal area was associated with a significantly higher recurrence rate of CSDH than placement at the frontal area (p=0.006) and the brain re-expansion rate (BRR) was much lower than placement at the frontal area (p<0.001). CONCLUSION: The operation may be delayed in high- and mixed-density groups, unless severe symptoms or signs are present. In addition, placing the catheter tip at the frontal area helps to reduce the incidence of postoperative recurrence of CSDH and to increase the BRR.

Molecular Detection and Seroprevalence of Babesia microti among Stock Farmers in Khutul City, Selenge Province, Mongolia

Babesiosis is an emerging tick-borne disease in humans worldwide; however, little is known about the frequency of infection or prevalence of this disease in other parts of the world, excluding North America. In this study, we aimed to investigate Babesia microti infection frequency in a human population in Mongolia. One hundred blood samples were collected from stock farmers living in Khutul city of Selenge province, Mongolia. The sera and DNA from blood samples were evaluated for the presence of B. microti infection by using indirect fluorescent antibody (IFA) tests and PCR. The positive detection rates obtained using the IFA tests and PCR assays were 7% and 3%, respectively. This study is the first to detect of B. microti infections based on antibody seroprevalence or PCR assays for the presence of B. microti DNA in a Mongolian population.

Staphylococcus Species in the Dental and Medical Environment

Staphylococcus species are one of prevalent pathogens found in hospitals. Microbes that are a primary cause of nosocomial infection were isolated from a dental and medical environment it may assist the reader to explain what this is and how it differs from the 'dental health care providers and ward health care providers'. To investigate the distribution of staphylococcus species in this environment, we used vitek II to measure drug sensitivity, and further performed biochemical testing. The isolation rate of staphylococcus species from the dental and medical environment was 100% but from dental health care providers and ward health care providers were 44.4% and 33.3%, respectively. In the analyses, staphylococcus species showed resistance to diffusion of cefoxitin and oxacillin discs. These staphylococci may be sufficiently positive for the mecA gene. Our results suggest that staphylococci might be an important cause of nosocomial infection in the dental clinic.