Coronavirus Activates an Altruistic Stem Cell–Mediated Defense Mechanism that Reactivates Dormant Tuberculosis
2021; Elsevier BV; Volume: 191; Issue: 7 Linguagem: Inglês
10.1016/j.ajpath.2021.03.011
ISSN1525-2191
AutoresLekhika Pathak, Sukanya Gayan, Bidisha Pal, Joyeeta Talukdar, Seema Bhuyan, Sorra Sandhya, Herman Yeger, Debabrat Baishya, Bikul Das,
Tópico(s)Neonatal Respiratory Health Research
ResumoWe postulate that similar to bacteria, adult stem cells may also exhibit an altruistic defense mechanism to protect their niche against external threat. Herein, we report mesenchymal stem cell (MSC)–based altruistic defense against a mouse model of coronavirus, murine hepatitis virus-1 (MHV-1) infection of lung. MHV-1 infection led to reprogramming of CD271+ MSCs in the lung to an enhanced stemness phenotype that exhibits altruistic behavior, as per previous work in human embryonic stem cells. The reprogrammed MSCs exhibited transient expansion for 2 weeks, followed by apoptosis and expression of stemness genes. The conditioned media of the reprogrammed MSCs exhibited direct antiviral activity in an in vitro model of MHV-1–induced toxicity to type II alveolar epithelial cells by increasing their survival/proliferation and decreasing viral load. Thus, the reprogrammed MSCs can be identified as altruistic stem cells (ASCs), which exert a unique altruistic defense against MHV-1. In a mouse model of MSC-mediated Mycobacterium tuberculosis (MTB) dormancy, MHV-1 infection in the lung exhibited 20-fold lower viral loads than the MTB-free control mice on the third week of viral infection, and exhibited six-fold increase of ASCs, thereby enhancing the altruistic defense. Notably, these ASCs exhibited intracellular replication of MTB, and their extracellular release. Animals showed tuberculosis reactivation, suggesting that dormant MTB may exploit ASCs for disease reactivation. We postulate that similar to bacteria, adult stem cells may also exhibit an altruistic defense mechanism to protect their niche against external threat. Herein, we report mesenchymal stem cell (MSC)–based altruistic defense against a mouse model of coronavirus, murine hepatitis virus-1 (MHV-1) infection of lung. MHV-1 infection led to reprogramming of CD271+ MSCs in the lung to an enhanced stemness phenotype that exhibits altruistic behavior, as per previous work in human embryonic stem cells. The reprogrammed MSCs exhibited transient expansion for 2 weeks, followed by apoptosis and expression of stemness genes. The conditioned media of the reprogrammed MSCs exhibited direct antiviral activity in an in vitro model of MHV-1–induced toxicity to type II alveolar epithelial cells by increasing their survival/proliferation and decreasing viral load. Thus, the reprogrammed MSCs can be identified as altruistic stem cells (ASCs), which exert a unique altruistic defense against MHV-1. In a mouse model of MSC-mediated Mycobacterium tuberculosis (MTB) dormancy, MHV-1 infection in the lung exhibited 20-fold lower viral loads than the MTB-free control mice on the third week of viral infection, and exhibited six-fold increase of ASCs, thereby enhancing the altruistic defense. Notably, these ASCs exhibited intracellular replication of MTB, and their extracellular release. Animals showed tuberculosis reactivation, suggesting that dormant MTB may exploit ASCs for disease reactivation. The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)–mediated coronavirus disease 2019 (COVID-19) pandemic demonstrates the ability of an emerging virus to generate chaos in our modern health care system and a severe strain on global economics. Postpandemic, the SARS-CoV-2 might activate dormant bacterial infections in the long term. As per prior history, tuberculosis is one of the key bacterial infections affected by viral pandemics.1Walaza S. Tempia S. Dawood H. Variava E. Wolter N. Dreyer A. Moyes J. Von Mollendorf C. McMorrow M. Von Gottberg A. Haffejee S. Venter M. Treurnicht F.K. Hellferscee O. Martinson N.A. Ismail N. Cohen C. 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Influenza virus infection is associated with increased risk of death amongst patients hospitalized with confirmed pulmonary tuberculosis in South Africa, 2010-2011.BMC Infect Dis. 2015; 15: 26Crossref PubMed Scopus (43) Google Scholar In the year 2009 during the influenza A (H1N1) pandemic, a worse prognosis of influenza was observed in patients with TB or multidrug-resistant TB.10Pandemic Influenza A (H1N1) 2009: Considerations for Tuberculosis Care Services, Paul Nunn (Coordinator) and Dennis Falzon (Medical Officer). Stop TB Department, World Health Organisation, Geneva, Switzerland2009Google Scholar,11Park Y. Chin B.S. Han S.H. Yun Y. Kim Y.J. Choi J.Y. Kim C.O. Song Y.G. Kim J.M. Pandemic influenza (H1N1) and Mycobacterium tuberculosis co-infection.Tuberc Respir Dis (Seoul). 2014; 76: 84-87Crossref PubMed Scopus (8) Google Scholar Interestingly, SARS-CoV-1 and Middle East respiratory syndrome coronavirus-infected patients were reported to develop pulmonary TB.4Low J.G. Lee C.C. Leo Y.S. Severe acute respiratory syndrome and pulmonary tuberculosis.Clin Infect Dis. 2004; 38: e123-e125Crossref PubMed Scopus (27) Google Scholar,5Alfaraj S.H. Al-Tawfiq J.A. Altuwaijri T.A. Memish Z.A. Middle east respiratory syndrome coronavirus and pulmonary tuberculosis coinfection: implications for infection control.Intervirology. 2017; 60: 53-55Crossref PubMed Scopus (39) Google Scholar In a mouse model, influenza A virus causes rapid development of pulmonary TB lesions12Volkert M. Pierce C. Horsfall F.L. Dubos R.J. The enhancing effect of concurrent infection with pneumotropic viruses on pulmonary tuberculosis in mice.J Exp Med. 1947; 86: 203-214Crossref PubMed Scopus (19) Google Scholar and an increase in the mycobacterium load in the liver.13Co D.O. Hogan L.H. Karman J. Heninger E. Vang S. Wells K. Kawaoka Y. Sandor M. Interactions between T cells responding to concurrent mycobacterial and influenza infections.J Immunol. 2006; 177: 8456-8465Crossref PubMed Scopus (16) Google Scholar Another mouse model of influenza A virus and Mycobacterium tuberculosis (MTB) co-infection leads to enhanced MTB growth by a type I interferon signaling pathway.6Redford P.S. Mayer-Barber K.D. McNab F.W. Stavropoulos E. Wack A. Sher A. O'Garra A. Influenza A virus impairs control of Mycobacterium tuberculosis coinfection through a type I interferon receptor-dependent pathway.J Infect Dis. 2014; 209: 270-274Crossref PubMed Scopus (94) Google Scholar However, severe inflammation in the lung is a common outcome of coronavirus infection,14Okabayashi T. Kariwa H. Yokota S. Iki S. Indoh T. Yokosawa N. Takashima I. Tsutsumi H. Fujii N. Cytokine regulation in SARS coronavirus infection compared to other respiratory virus infections.J Med Virol. 2006; 78: 417-424Crossref PubMed Scopus (118) Google Scholar a symptom that is also commonly observed in TB patients.15Kaufmann S.H. Dorhoi A. Inflammation in tuberculosis: interactions, imbalances and interventions.Curr Opin Immunol. 2013; 25: 441-449Crossref PubMed Scopus (92) Google Scholar Thus, it is possible that coronavirus infection-causing inflammation may also reactivate dormant MTB (dMTB) in the lung, which has not yet been studied. Several studies have investigated TB dormancy in the adult stem cell niches.16Beamer G. Major S. Das B. Campos-Neto A. Bone marrow mesenchymal stem cells provide an antibiotic-protective niche for persistent viable Mycobacterium tuberculosis that survive antibiotic treatment.Am J Pathol. 2014; 184: 3170-3175Abstract Full Text Full Text PDF PubMed Scopus (33) Google Scholar, 17Das B. Kashino S.S. Pulu I. Kalita D. Swami V. Yeger H. Felsher D.W. Campos-Neto A. CD271(+) bone marrow mesenchymal stem cells may provide a niche for dormant Mycobacterium tuberculosis.Sci Transl Med. 2013; 5: 170-170ra13Crossref Scopus (147) Google Scholar, 18Garhyan J. Bhuyan S. Pulu I. Kalita D. Das B. Bhatnagar R. Preclinical and clinical evidence of Mycobacterium tuberculosis persistence in the hypoxic niche of bone marrow mesenchymal stem cells after therapy.Am J Pathol. 2015; 185: 1924-1934Abstract Full Text Full Text PDF PubMed Scopus (35) Google Scholar These stem cells reside in the bone marrow (BM) niche19Jones E. McGonagle D. Human bone marrow mesenchymal stem cells in vivo.Rheumatology (Oxford). 2008; 47: 126-131Crossref PubMed Scopus (231) Google Scholar and in the area of inflammation.20Spaeth E.L. Kidd S. Marini F.C. Tracking inflammation-induced mobilization of mesenchymal stem cells.Methods Mol Biol. 2012; 904: 173-190PubMed Google Scholar A rare fraction of cells in the BM, the CD271+ BM–mesenchymal stem cells (CD271+ BM-MSCs) has been identified as the potential niche for dMTB in mice and in successfully treated TB patients.17Das B. Kashino S.S. Pulu I. Kalita D. Swami V. Yeger H. Felsher D.W. Campos-Neto A. CD271(+) bone marrow mesenchymal stem cells may provide a niche for dormant Mycobacterium tuberculosis.Sci Transl Med. 2013; 5: 170-170ra13Crossref Scopus (147) Google Scholar In this stem cell niche, MTB remains dormant, maintaining reactivation potential. More importantly, we have developed a mouse model of stem cell–mediated MTB dormancy.17Das B. Kashino S.S. Pulu I. Kalita D. Swami V. Yeger H. Felsher D.W. Campos-Neto A. CD271(+) bone marrow mesenchymal stem cells may provide a niche for dormant Mycobacterium tuberculosis.Sci Transl Med. 2013; 5: 170-170ra13Crossref Scopus (147) Google Scholar Briefly, streptomycin-dependent mutant 18b strain-infected mice exhibit lung infection following 3 weeks of streptomycin treatment. These mice develop granulomas, and acquire humoral immunity against the bacteria. Following streptomycin starvation for 6 months, the bacteria acquire a nonreplicating status. These bacteria can primarily be detected in the CD271+ MSCs of BM, but a few are also present in the CD271+ MSCs of lung.17Das B. Kashino S.S. Pulu I. Kalita D. Swami V. Yeger H. Felsher D.W. Campos-Neto A. CD271(+) bone marrow mesenchymal stem cells may provide a niche for dormant Mycobacterium tuberculosis.Sci Transl Med. 2013; 5: 170-170ra13Crossref Scopus (147) Google Scholar Furthermore, MTB harboring CD271+ MSCs reside in the hypoxic niche of BM.18Garhyan J. Bhuyan S. Pulu I. Kalita D. Das B. Bhatnagar R. Preclinical and clinical evidence of Mycobacterium tuberculosis persistence in the hypoxic niche of bone marrow mesenchymal stem cells after therapy.Am J Pathol. 2015; 185: 1924-1934Abstract Full Text Full Text PDF PubMed Scopus (35) Google Scholar Notably, in this model, the significant increase of MTB–colony-forming units (CFUs) in the non–CD271+ MSC compartment of lung can be used as a sign of MTB reactivation. Additionally, the potential reprogramming of the CD271+ MSCs to enhanced stemness phenotype having altruistic behavior21BD Altruistic stem cells and cancer stem cells.in: Rajasekhar V.K. Cancer Stem Cells. John Wiley & Sons, Hoboken, NJ2014: 89-106Google Scholar,22Das B. Bayat-Mokhtari R. Tsui M. Lotfi S. Tsuchida R. Felsher D.W. Yeger H. HIF-2alpha suppresses p53 to enhance the stemness and regenerative potential of human embryonic stem cells.Stem Cells. 2012; 30: 1685-1695Crossref PubMed Scopus (57) Google Scholar can also be studied in this model. Briefly, the enhanced stemness is a transient phenotype of stem cells characterized by their ability to not only maintains stemness, but also secrete cytoprotective agents during extreme oxidative stress or inflammation.21BD Altruistic stem cells and cancer stem cells.in: Rajasekhar V.K. Cancer Stem Cells. John Wiley & Sons, Hoboken, NJ2014: 89-106Google Scholar,22Das B. Bayat-Mokhtari R. Tsui M. Lotfi S. Tsuchida R. Felsher D.W. Yeger H. HIF-2alpha suppresses p53 to enhance the stemness and regenerative potential of human embryonic stem cells.Stem Cells. 2012; 30: 1685-1695Crossref PubMed Scopus (57) Google Scholar The enhanced stemness phenotype was characterized by exposing human embryonic stem (ES) cells to the microenvironment of oxidative stress. In such hostile microenvironment, some of the ES cells underwent enhanced stemness reprogramming by activating a hypoxia-inducible factor (HIF)-2α stemness pathway that altered the p53/mouse double minute 2 homolog oscillation to a transient state of low p53. The low p53 state permits these cells to maintain a state of enhanced stemness (state of self-renewal and self-sufficiency) in the microenvironment oxidative stress.22Das B. Bayat-Mokhtari R. Tsui M. Lotfi S. Tsuchida R. Felsher D.W. Yeger H. HIF-2alpha suppresses p53 to enhance the stemness and regenerative potential of human embryonic stem cells.Stem Cells. 2012; 30: 1685-1695Crossref PubMed Scopus (57) Google Scholar While in such hostile microenvironment, naive stem cells usually undergo differentiation/apoptosis,21BD Altruistic stem cells and cancer stem cells.in: Rajasekhar V.K. Cancer Stem Cells. John Wiley & Sons, Hoboken, NJ2014: 89-106Google Scholar and the reprogrammed ES cells not only maintain stemness, but also self-renew. Thus, these reprogrammed ES cells gain fitness. However, instead of becoming the dominant subpopulation, these cells sacrifice their self-fitness to enhance the fitness of neighboring cells under stress by secreting glutathione, an intracellular antioxidant that cells do not actively secrete. Moreover, the cultured supernatant of these reprogramed ES cells defend hematopoietic and mesenchymal stem cells from oxidative stress–induced differentiation/apoptosis. This is considered an altruistic behavior, a form of stem cell altruism,23Das B. The Science Behind Squalene: The Human Antioxidant. Toronto Medical Pub., for the International Council for Bionutrient Research, Toronto, Canada2000: 187Google Scholar as these glutathione-secreting ES cells sacrifice their newly acquired fitness by returning to basal state of p53/MDM2 oscillation, leading to p53-mediated apoptosis/differentiation.22Das B. Bayat-Mokhtari R. Tsui M. Lotfi S. Tsuchida R. Felsher D.W. Yeger H. HIF-2alpha suppresses p53 to enhance the stemness and regenerative potential of human embryonic stem cells.Stem Cells. 2012; 30: 1685-1695Crossref PubMed Scopus (57) Google Scholar Hence, these transient stem cells acquiring enhanced stemness phenotype can be termed as altruistic stem cells (ASCs) as opposed to competitive stem cells, which eliminate weak neighbors during stress.21BD Altruistic stem cells and cancer stem cells.in: Rajasekhar V.K. Cancer Stem Cells. John Wiley & Sons, Hoboken, NJ2014: 89-106Google Scholar,24Talukdar J.B.R. Garhyan J. Pal B. Sandhya S. Gayan S. Sarma A. Mokhtari R.B. Li H. Phukan J. 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Griffin A.S. Gardner A. Diggle S.P. Social evolution theory for microorganisms.Nat Rev Microbiol. 2006; 4: 597-607Crossref PubMed Scopus (827) Google Scholar ASC mechanisms may also have evolved as a potent stem cell defense of cytoprotection from stressful microenvironment-induced differentiation/apoptosis.21BD Altruistic stem cells and cancer stem cells.in: Rajasekhar V.K. Cancer Stem Cells. John Wiley & Sons, Hoboken, NJ2014: 89-106Google Scholar Developing an experimental model to study ASC-based defense will strengthen our growing understanding of stem cell niche defense against pathogen invasion, and provide novel insight about how pathogens, such as dMTB, may hijack this defense for their own benefit. Herein, we hypothesize that following viral infection, lung MSCs present in the alveolar stem cell niche28Lee J.W. Fang X. Krasnodembskaya A. Howard J.P. Matthay M.A. Concise review: mesenchymal stem cells for acute lung injury: role of paracrine soluble factors.Stem Cells. 2011; 29: 913-919Crossref PubMed Scopus (333) Google Scholar,29Zhu H. Xiong Y. Xia Y. Zhang R. Tian D. Wang T. Dai J. Wang L. Yao H. Jiang H. Yang K. Liu E. Shi Y. Fu Z. Gao L. Zou L. Therapeutic effects of human umbilical cord-derived mesenchymal stem cells in acute lung injury mice.Sci Rep. 2017; 7: 39889Crossref PubMed Scopus (69) Google Scholar may also exhibit altruistic defense by reprogramming to ASCs. The dormant MTB hiding in the intracellular compartment of these MSCs may then exploit the ASC reprograming for TB reactivation.25Pathak L. Das B. Initiation of post-primary tuberculosis of the lungs: exploring the secret role of bone marrow derived stem cells.Front Immunol. 2021; 11: 594572Crossref PubMed Scopus (6) Google Scholar Therefore, this mouse model of stem cell–mediated MTB dormancy was used to find out if coronavirus can reactivate dMTB by inducing MSC to ASC reprogramming. A mouse coronavirus strain, the murine hepatitis virus-1 (MHV-1), that represents clinically relevant human coronavirus SARS-CoV-1 infection, was used.30De Albuquerque N. Baig E. Ma X. Zhang J. He W. Rowe A. Habal M. Liu M. Shalev I. Downey G.P. Gorczynski R. Butany J. Leibowitz J. Weiss S.R. McGilvray I.D. Phillips M.J. Fish E.N. Levy G.A. Murine hepatitis virus strain 1 produces a clinically relevant model of severe acute respiratory syndrome in A/J mice.J Virol. 2006; 80: 10382-10394Crossref PubMed Scopus (117) Google Scholar, 31Khanolkar A. Fulton R.B. Epping L.L. Pham N.L. Tifrea D. Varga S.M. Harty J.T. T cell epitope specificity and pathogenesis of mouse hepatitis virus-1-induced disease in susceptible and resistant hosts.J Immunol. 2010; 185: 1132-1141Crossref PubMed Scopus (16) Google Scholar, 32Khanolkar A. Hartwig S.M. Haag B.A. Meyerholz D.K. Harty J.T. Varga S.M. Toll-like receptor 4 deficiency increases disease and mortality after mouse hepatitis virus type 1 infection of susceptible C3H mice.J Virol. 2009; 83: 8946-8956Crossref PubMed Scopus (45) Google Scholar MHV-1 infection causes acute lung inflammation by inducing acute respiratory infection within 2 to 4 days in C57BL/6 mice by increasing viral load. Animals exhibit an elevated level of proinflammatory cytokines, such as tumor necrosis factor (TNF)-α, during 2 to 14 days postinfection30De Albuquerque N. Baig E. Ma X. Zhang J. He W. Rowe A. Habal M. Liu M. Shalev I. Downey G.P. Gorczynski R. Butany J. Leibowitz J. Weiss S.R. McGilvray I.D. Phillips M.J. Fish E.N. Levy G.A. Murine hepatitis virus strain 1 produces a clinically relevant model of severe acute respiratory syndrome in A/J mice.J Virol. 2006; 80: 10382-10394Crossref PubMed Scopus (117) Google Scholar,33Khanolkar A. Hartwig S.M. Haag B.A. Meyerholz D.K. Epping L.L. Haring J.S. Varga S.M. Harty J.T. Protective and pathologic roles of the immune response to mouse hepatitis virus type 1: implications for severe acute respiratory syndrome.J Virol. 2009; 83: 9258-9272Crossref PubMed Scopus (35) Google Scholar and then fully recover.30De Albuquerque N. Baig E. Ma X. Zhang J. He W. Rowe A. Habal M. Liu M. Shalev I. Downey G.P. Gorczynski R. Butany J. Leibowitz J. Weiss S.R. McGilvray I.D. Phillips M.J. Fish E.N. Levy G.A. Murine hepatitis virus strain 1 produces a clinically relevant model of severe acute respiratory syndrome in A/J mice.J Virol. 2006; 80: 10382-10394Crossref PubMed Scopus (117) Google Scholar Therefore, the MHV-1–infected C57BL/6 mice can be utilized for MTB reactivation study. Herein, MHV-1 infection was shown to cause dMTB reactivation in the mouse model of stem cell–mediated MTB dormancy. Furthermore, MHV-1 was found to reprogram the CD271+ MSCs to an enhanced stemness or ASC phenotype, and culture supernatant of these reprogrammed MSCs was found to enhance the survival/proliferation of MHV-1–infected type II alveolar epithelial (ATII) cells. After 2 weeks of expansion in the MHV-1–infected lung, these reprogrammed MSCs activated p53 upstream genes involved in apoptosis/differentiation, and underwent apoptosis, thus sacrificing self-fitness while enhancing the fitness of the alveolar epithelial cells, a behavior akin to ES cells exhibiting enhanced stemness or ASC phenotype. Thus, these findings may facilitate the study of proposed ASC-mediated altruistic defense of stem cell niche.21BD Altruistic stem cells and cancer stem cells.in: Rajasekhar V.K. Cancer Stem Cells. John Wiley & Sons, Hoboken, NJ2014: 89-106Google Scholar,25Pathak L. Das B. Initiation of post-primary tuberculosis of the lungs: exploring the secret role of bone marrow derived stem cells.Front Immunol. 2021; 11: 594572Crossref PubMed Scopus (6) Google Scholar,34Pal B. Das B. In vitro culture of naive human bone marrow mesenchymal stem cells: a stemness based approach.Front Cell Dev Biol. 2017; 5: 69Crossref PubMed Scopus (33) Google Scholar The remarkable significance of this study is that although MHV-1 infection causes dMTB reactivation, it also activates an ASC-based innate defense mechanism against the virus and that could be further explored to develop therapeutics to target coronavirus. All of the necessary experimental procedures were approved and undertaken inside Biosafety Cabinet class II facility in accordance with guidelines of Institutional Bio-safety Committee of KaviKrishna Laboratory. Streptomycin-auxotrophic mutant MTB strain18b (gifted by Prof. Stewart T. Cole, Ecole Polytechinque Federale de Lausanne, Lausanne, Switzerland) was cultured in BBL Middlebrook 7H9 broth with glycerol (BD Biosciences, Gurugram, Haryana, India; number 221832) along with 50 μg/mL of streptomycin sulfate. It was maintained at 37°C and 5% CO2 with occasional shaking until the midlogarithmic phase was reached, OD approximating to 1. All of the necessary experimental procedures were undertaken in accordance with approvals of Institutional Animal Ethics Committee, Gauhati University, and Institutional Ethics Committee, KaviKrishna Laboratory. The 6- to 8-week–old C57BL/6 female mice were obtained from National Institution of Nutrition (Hyderabad, India) and were maintained in the animal house of Gauhati University at pathogen-free condition, as previously described.17Das B. Kashino S.S. Pulu I. Kalita D. Swami V. Yeger H. Felsher D.W. Campos-Neto A. CD271(+) bone marrow mesenchymal stem cells may provide a niche for dormant Mycobacterium tuberculosis.Sci Transl Med. 2013; 5: 170-170ra13Crossref Scopus (147) Google Scholar The mouse model of MTB dormancy was developed in 6 months. Briefly, streptomycin-auxotrophic mutant MTB strain 18b cell suspension was prepared in phosphate-buffered saline–Tween 80 (0.05%), sonicated for 15 seconds, and intravenously injected with 2 × 106 CFUs per mouse. Initially for 3 weeks, streptomycin was administered (3 mg/mouse in 200 μL of normal saline) daily for establishing infection. Then, no streptomycin treatment was provided for 6 months to establish bacterial dormancy. Following 6 months of streptomycin starvation, lung tissue was dissociated; CD271+/CD45– MSCs and non-CD271+ cells were isolated by magnetic sorting, as previously explained.18Garhyan J. Bhuyan S. Pulu I. Kalita D. Das B. Bhatnagar R. Preclinical and clinical evidence of Mycobacterium tuberculosis persistence in the hypoxic niche of bone marrow mesenchymal stem cells after therapy.Am J Pathol. 2015; 185: 1924-1934Abstract Full Text Full Text PDF PubMed Scopus (35) Google Scholar The magnetically sorted CD271+ MSCs and non-CD271+ cells were subjected to MTB-CFU assay. Consistent with previous findings,17Das B. Kashino S.S. Pulu I. Kalita D. Swami V. Yeger H. Felsher D.W. Campos-Neto A. CD271(+) bone marrow mesenchymal stem cells may provide a niche for dormant Mycobacterium tuberculosis.Sci Transl Med. 2013; 5: 170-170ra13Crossref Scopus (147) Google Scholar a small number of MTB-CFUs (38 ± 10 MTB-CFU/lung) were obtained only in CD271+ MSCs. Thus, mouse model of MTB dormancy was developed. Streptomycin was injected intraperitoneally (3 mg/mouse in 200 μL of normal saline) daily for 3 weeks17Das B. Kashino S.S. Pulu I. Kalita D. Swami V. Yeger H. Felsher D.W. Campos-Neto A. CD271(+) bone marrow mesenchymal stem cells may provide a niche for dormant Mycobacterium tuberculosis.Sci Transl Med. 2013; 5: 170-170ra13Crossref Scopus (147) Google Scholar with or without MHV-1 infection30De Albuquerque N. Baig E. Ma X. Zhang J. He W. Rowe A. Habal M. Liu M. Shalev I. Downey G.P. Gorczynski R. Butany J. Leibowitz J. Weiss S.R. McGilvray I.D. Phillips M.J. Fish E.N. Levy G.A. Murine hepatitis virus strain 1 produces a clinically relevant model of severe acute respiratory syndrome in A/J mice.J Virol. 2006; 80: 10382-10394Crossref PubMed Scopus (117) Google Scholar,32Khanolkar A. Hartwig S.M. Haag B.A. Meyerholz D.K. Harty J.T. Varga S.M. Toll-like receptor 4 deficiency increases disease and mortality after mouse hepatitis virus type 1 infection of susceptible C3H mice.J Virol. 2009; 83: 8946-8956Crossref PubMed Scopus (45) Google Scholar to develop a mouse model of reactivation. In a separate experiment, streptomycin-treated mice were also treated with immunosuppressive agents dexamethasone (0.08 mg/day, 6 times a week) or amino guanidine (2.5% wt/vol in drinking water) to cause dMTB reactivation.35Scanga C.A. Mohan V.P. Joseph H. Yu K. Chan J. Flynn J.L. Reactivation of latent tuberculosis: variations on the Cornell murine model.Infect Immun. 1999; 67: 4531-4538Crossref PubMed Google Scholar The treated mice were observed for stipulated period of time, and then sacrificed to collect lung tissues. The MTB-CFU of lung was evaluated for MTB reactivation. The mouse lung tissues were dissociated using collagenase/lipase and then cells were subjected to magnetic sorting to isolate CD271+ MSCs, as explained previously.17Das B. Kashino S.S. Pulu I. Kalita D. Swami V. Yeger H. Felsher D.W. Campos-Neto A. CD271(+) bone marrow mesenchymal stem cells may provide a niche for dormant Mycobacterium tuberculosis.Sci Transl Med. 2013; 5: 170-170ra13Crossref Scopus (147) Google Scholar,18Garhyan J. Bhuyan S. Pulu I. Kalita D. Das B. Bhatnagar R. Preclinical and clinical evidence of Mycobacterium tuberculosis persistence in the hypoxic niche of bone marrow mesenchymal stem cells after therapy.Am J Pathol. 2015; 185: 1924-1934Abstract Full Text Full Text PDF PubMed Scopus (35) Google Scholar Briefly, CD45– cell
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