Human Granuloma In Vitro Model, for TB Dormancy and Resuscitation

Abstract

Tuberculosis (TB) is responsible for death of nearly two million people in the world annually. Upon infection, Mycobacterium tuberculosis (Mtb) causes formation of granuloma where the pathogen goes into dormant state and can live for decades before resuscitation to develop active disease when the immune system of the host is weakened and/or suppressed. In an attempt to better understand host-pathogen interactions, several groups have been developing in vitro models of human tuberculosis granuloma. However, to date, an in vitro granuloma model in which Mtb goes into dormancy and can subsequently resuscitate under conditions that mimic weakening of the immune system has not been reported. We describe the development of a biomimetic in vitro model of human tuberculosis granuloma using human primary leukocytes, in which the Mtb exhibited characteristics of dormant mycobacteria as demonstrated by (1) loss of acid-fastness, (2) accumulation of lipid bodies (3) development of rifampicin-tolerance and (4) gene expression changes. Further, when these micro granulomas were treated with immunosuppressant anti-tumor necrosis factor-alpha monoclonal antibodies (anti-TNFα mAbs), resuscitation of Mtb was observed as has been found in humans. In this human in vitro granuloma model triacylglycerol synthase 1deletion mutant (Δtgs1) with impaired ability to accumulate triacylglycerides (TG), but not the complemented mutant, could not go into dormancy. Deletion mutant of lipY, with compromised ability to mobilize the stored TG, but not the complemented mutant, was unable to come out of dormancy upon treatment with anti-TNFα mAbs. In conclusion, we have developed an in vitro human tuberculosis granuloma model that largely exhibits functional features of dormancy and resuscitation observed in human tuberculosis.

Rv2744c Is a PspA Ortholog That Regulates Lipid Droplet Homeostasis and Nonreplicating Persistence in Mycobacterium tuberculosis

“…Consistent with this observation, Mycobacterium bovis BCG strains that are inhibited in their ability to mediate TAG hydrolysis are attenuated for survival in vitro during and following resuscitation from NRP (60,61). Similarly, an M. tuberculosis mutant deleted in triacylglycerol-1 synthase (⌬tgs1), an enzyme used to make TAGs, is compromised in its ability to accumulate intracellular TAGs and is unable to enter into NRP in an in vitro human TB granuloma model (73). Finally, an M. tuberculosis mutant deleted in lipase lipY, which is required for TAG hydrolysis, is compromised in its ability to mobilize stored TAGs and is unable to reactivate from NRP in the granuloma model system (73).…”

“…Similarly, an M. tuberculosis mutant deleted in triacylglycerol-1 synthase (⌬tgs1), an enzyme used to make TAGs, is compromised in its ability to accumulate intracellular TAGs and is unable to enter into NRP in an in vitro human TB granuloma model (73). Finally, an M. tuberculosis mutant deleted in lipase lipY, which is required for TAG hydrolysis, is compromised in its ability to mobilize stored TAGs and is unable to reactivate from NRP in the granuloma model system (73). Thus, LDs represent an important energy reservoir utilized by M. tuberculosis for survival during and resuscitation from NRP.…”

A 3D Human Lung Tissue Model for Functional Studies on <em>Mycobacterium tuberculosis</em> Infection

“…Single cell cultures of human macrophages or other peripheral blood cells have often been used 6,7 . The disadvantage of this approach is that they cannot reflect the dynamics of different cell types operating together in a lung tissue exposed to M. tuberculosis.…”

Detection of Mycobacterium Tuberculosis among Infertile Patients Suspected with Female Genital Tuberculosis

“…A positive chest X-ray for healed or active pulmonary tuberculosis, contact history, elevated erythrocyte sedimentation rate (ESR), positive tuberculin test and sampling by laparoscopy may specify the need for further investigations [19,20,21]. Studies such as serial sections of tissues need to be studied, because the lesions are frequently erratic and that a positive endometrial culture for TB, is found only in about 25% of cases of tuberculous endometritis as the endometrium is often focal and the functionalis layer is shed every four weeks (granulomas take two weeks to develop) [22,23]. Moreover, due to the cyclical shedding of the endometrium, granulomas do not have enough time to form, so the granulomatous endometrium may not show evidence of tuberculosis in all the cycles.…”