scholarly journals Evidence of the Extrahepatic Replication of Hepatitis E Virus in Human Endometrial Stromal Cells

Pathogens ◽  
2020 ◽  
Vol 9 (4) ◽  
pp. 295 ◽  
Author(s):  
Mohamed A. El-Mokhtar ◽  
Essam R. Othman ◽  
Maha Y. Khashbah ◽  
Ali Ismael ◽  
Mohamed AA Ghaliony ◽  
...  
Keyword(s):  
Capsid Protein ◽  
Stromal Cells ◽  
Hepatitis E Virus ◽  
Inflammatory Responses ◽  
Hepatitis E ◽  
Endometrial Stromal Cells ◽  
Extrahepatic Replication ◽  
Efficient Replication ◽  
Over Time

Hepatitis E virus (HEV) is the most common cause of acute viral hepatitis worldwide. The tropism of HEV is not restricted to the liver, and the virus replicates in other organs. Not all the extrahepatic targets for HEV are identified. Herein, we found that non-decidualized primary human endometrial stromal cells (PHESCs), which are precursors for the decidua and placenta, are susceptible to HEV infection. PHESCs, isolated from healthy non-pregnant women (n = 5), were challenged with stool-derived HEV-1 and HEV-3. HEV RNA was measured by qPCR, and HEV capsid protein was assessed by flow cytometry, immunofluorescence (IF), and ELISA. HEV infection was successfully established in PHESCs. Intracellular and extracellular HEV RNA loads were increased over time, indicating efficient replication in vitro. In addition, HEV capsid protein was detected intracellularly in the HEV-infected PHESCs and accumulated extracellularly over time, confirming the viral assembly and release from the infected cells. HEV-1 replicated more efficiently in PHESCs than HEV-3 and induced more inflammatory responses. Ribavirin (RBV) treatment abolished the replication of HEV in PHESCs. In conclusion, PHESCs are permissive to HEV infection and these cells could be an endogenous source of HEV infection during pregnancy and mediate HEV vertical transmission.

scholarly journals Replication of Hepatitis E Virus (HEV) in Primary Human-Derived Monocytes and Macrophages In Vitro

Vaccines ◽  
2020 ◽  
Vol 8 (2) ◽  
pp. 239 ◽  
Author(s):  
Ibrahim M. Sayed ◽  
Mohamed Ismail Seddik ◽  
Marwa A. Gaber ◽  
Saber H. Saber ◽  
Sahar A. Mandour ◽  
...  
Keyword(s):  
Capsid Protein ◽  
Inflammatory Responses ◽  
Organ Transplant ◽  
Immunocompromised Patients ◽  
Replicative Form ◽  
Extrahepatic Manifestations ◽  
Transplant Patients ◽  
Infected Cells ◽  
Over Time

HEV is the most causative agent of acute viral hepatitis globally. HEV causes acute, chronic, and extrahepatic manifestations. Chronic HEV infection develops in immunocompromised patients such as organ transplant patients, HIV-infected patients, and leukemic patients. The source of chronic HEV infection is not known. Also, the source of extrahepatic manifestations associated with HEV infection is still unclear. Hepatotropic viruses such as HCV and HBV replicate in peripheral blood mononuclear cells (PBMCs) and these cells become a source of chronic reactivation of the infections in allograft organ transplant patients. Herein, we reported that PBMCs and bone marrow-derived macrophages (BMDMs), isolated from healthy donors (n = 3), are susceptible to HEV in vitro. Human monocytes (HMOs), human macrophages (HMACs), and human BMDMs were challenged with HEV-1 and HEV-3 viruses. HEV RNA was measured by qPCR, the marker of the intermediate replicative form (ds-RNA) was assessed by immunofluorescence, and HEV capsid protein was assessed by flow cytometry and ELISA. HEV infection was successfully established in primary HMOs, HMACs, and human BMDMs, but not in the corresponding cells of murine origin. Intermediate replicative form (ds RNA) was detected in HMOs and HMACs challenged with HEV. The HEV load was increased over time, and the HEV capsid protein was detected intracellularly in the HEV-infected cells and accumulated extracellularly over time, confirming that HEV completes the life cycle inside these cells. The HEV particles produced from the infected BMDMs were infectious to naive HMOs in vitro. The HEV viral load was comparable in HEV-1- and HEV-3-infected cells, but HEV-1 induced more inflammatory responses. In conclusion, HMOs, HMACs, and human BMDMs are permissive to HEV infection and these cells could be the source of chronic and recurrent infection, especially in immunocompromised patients. Replication of HEV in human BMDMs could be related to hematological disorders associated with extrahepatic manifestations.

Characterization of capsid protein (p495) of hepatitis E virus expressed in Escherichia coli and assembling into particles in vitro

Vaccine ◽  
2018 ◽  
Vol 36 (16) ◽  
pp. 2104-2111 ◽  
Author(s):  
Minghua Zheng ◽  
Jie Jiang ◽  
Xiao Zhang ◽  
Nan Wang ◽  
Kaihang Wang ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Capsid Protein ◽  
Hepatitis E Virus ◽  
Hepatitis E

scholarly journals Characterization of a Cell Culture System of Persistent Hepatitis E Virus Infection in the Human HepaRG Hepatic Cell Line

Viruses ◽  
2021 ◽  
Vol 13 (3) ◽  
pp. 406
Author(s):  
Marie Pellerin ◽  
Edouard Hirchaud ◽  
Yannick Blanchard ◽  
Nicole Pavio ◽  
Virginie Doceul
Keyword(s):  
Cell Culture ◽  
Culture System ◽  
Hepatitis E Virus ◽  
Hepatitis E ◽  
Inhibitory Effect ◽  
Cell Culture System ◽  
Efficient Replication ◽  
A Cell ◽  
Vitro Model

Hepatitis E virus (HEV) is considered as an emerging global health problem. In most cases, hepatitis E is a self-limiting disease and the virus is cleared spontaneously without the need of antiviral therapy. However, immunocompromised individuals can develop chronic infection and liver fibrosis that can progress rapidly to cirrhosis and liver failure. The lack of efficient and relevant cell culture system and animal models has limited our understanding of the biology of HEV and the development of effective drugs for chronic cases. In the present study, we developed a model of persistent HEV infection in human hepatocytes in which HEV replicates efficiently. This HEV cell culture system is based on differentiated HepaRG cells infected with an isolate of HEV-3 derived from a patient suffering from acute hepatitis E. Efficient replication was maintained for several weeks to several months as well as after seven successive passages on HepaRG naïve cells. Moreover, after six passages onto HepaRG, we found that the virus was still infectious after oral inoculation into pigs. We also showed that ribavirin had an inhibitory effect on HEV replication in HepaRG. In conclusion, this system represents a relevant and efficient in vitro model of HEV replication that could be useful to study HEV biology and identify effective antiviral drugs against chronic HEV infection.

scholarly journals Mutations within Potential Glycosylation Sites in the Capsid Protein of Hepatitis E Virus Prevent the Formation of Infectious Virus Particles

2007 ◽  
Vol 82 (3) ◽  
pp. 1185-1194 ◽  
Author(s):  
Judith Graff ◽  
Yi-Hua Zhou ◽  
Udana Torian ◽  
Hanh Nguyen ◽  
Marisa St. Claire ◽  
...  
Keyword(s):  
Cell Culture ◽  
Capsid Protein ◽  
Hepatitis E Virus ◽  
Signal Sequence ◽  
Rhesus Macaques ◽  
Hepatitis E ◽  
Velocity Sedimentation ◽  
Genome Replication ◽  
Glycosylation Sites

ABSTRACT Hepatitis E virus is a nonenveloped RNA virus. However, the single capsid protein resembles a typical glycoprotein in that it contains a signal sequence and potential glycosylation sites that are utilized when recombinant capsid protein is overexpressed in cell culture. In order to determine whether these unexpected observations were biologically relevant or were artifacts of overexpression, we analyzed capsid protein produced during a normal viral replication cycle. In vitro transcripts from an infectious cDNA clone mutated to eliminate potential glycosylation sites were transfected into cultured Huh-7 cells and into the livers of rhesus macaques. The mutations did not detectably affect genome replication or capsid protein synthesis in cell culture. However, none of the mutants infected rhesus macaques. Velocity sedimentation analyses of transfected cell lysates revealed that mutation of the first two glycosylation sites prevented virion assembly, whereas mutation of the third site permitted particle formation and RNA encapsidation, but the particles were not infectious. However, conservative mutations that did not destroy glycosylation motifs also prevented infection. Overall, the data suggested that the mutations were lethal because they perturbed protein structure rather than because they eliminated glycosylation.

scholarly journals Endometrial regeneration with endometrial epithelium: homologous orchestration with endometrial stroma as a feeder

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Ryo Yokomizo ◽  
Yukiko Fujiki ◽  
Harue Kishigami ◽  
Hiroshi Kishi ◽  
Tohru Kiyono ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Stromal Cells ◽  
Therapeutic Option ◽  
Three Dimensional ◽  
Fertility Treatment ◽  
Feeder Cells ◽  
Endometrial Stromal Cells ◽  
Thin Endometrium ◽  
Endometrial Epithelial Cells

Abstract Background Thin endometrium adversely affects reproductive success rates with fertility treatment. Autologous transplantation of exogenously prepared endometrium can be a promising therapeutic option for thin endometrium; however, endometrial epithelial cells have limited expansion potential, which needs to be overcome in order to make regenerative medicine a therapeutic strategy for refractory thin endometrium. Here, we aimed to perform long-term culture of endometrial epithelial cells in vitro. Methods We prepared primary human endometrial epithelial cells and endometrial stromal cells and investigated whether endometrial stromal cells and human embryonic stem cell-derived feeder cells could support proliferation of endometrial epithelial cells. We also investigated whether three-dimensional culture can be achieved using thawed endometrial epithelial cells and endometrial stromal cells. Results Co-cultivation with the feeder cells dramatically increased the proliferation rate of the endometrial epithelial cells. We serially passaged the endometrial epithelial cells on mouse embryonic fibroblasts up to passage 6 for 4 months. Among the human-derived feeder cells, endometrial stromal cells exhibited the best feeder activity for proliferation of the endometrial epithelial cells. We continued to propagate the endometrial epithelial cells on endometrial stromal cells up to passage 5 for 81 days. Furthermore, endometrial epithelium and stroma, after the freeze-thaw procedure and sequential culture, were able to establish an endometrial three-dimensional model. Conclusions We herein established a model of in vitro cultured endometrium as a potential therapeutic option for refractory thin endometrium. The three-dimensional culture model with endometrial epithelial and stromal cell orchestration via cytokines, membrane-bound molecules, extracellular matrices, and gap junction will provide a new framework for exploring the mechanisms underlying the phenomenon of implantation. Additionally, modified embryo culture, so-called “in vitro implantation”, will be possible therapeutic approaches in fertility treatment.

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