Detection of Mycobacterium tuberculosis Complex Using the Xpert MTB/RIF Ultra Assay on the Stool of Pediatric Patients in Dushanbe, Tajikistan.

We report the findings of a prospective laboratory diagnostic accuracy study to evaluate the sensitivity, specificity, and predictive values of the Xpert MTB/RIF Ultra assay for Mycobacterium tuberculosis detection in fresh stool specimens from children under 15 years of age with confirmed tuberculosis (TB) disease from Dushanbe, Tajikistan. Six hundred eighty-eight (688) participants were enrolled from April 2019 to October 2021. We identified 16 participants (2.3%) with confirmed TB disease, defined as ≥1 TB sign/symptom plus microbiologic confirmation. With the Xpert MTB/RIF Ultra assay for stool, we found a sensitivity of 68.8% (95% CI, 46.0 to 91.5) and a specificity of 98.7% (95% CI, 97.8 to 99.5) in confirmed TB disease. Our results are comparable to other published studies; however, our cohort was larger and our confirmed TB disease rate lower than most. We also demonstrated that this assay was feasible to implement in a centralized hospital laboratory in a low-middle-income Central Asian country. However, we encountered obstacles such as lack of staffing, material ruptures, outdated government protocols, and decreased case presentation due to COVID-19. We found eight patients whose only positive test was an Xpert Ultra stool assay. None needed treatment during the study; however, three were treated later, suggesting such cases require close observation. Our report is the first from Central Asia and one of a few from a low-middle-income country. We believe our study demonstrates the generalizability of the Xpert MTB/RIF Ultra assay on fresh stool specimens from children and provides further evidence supporting WHO's approval of this diagnostic strategy. IMPORTANCE The importance of this report is that it provides further support for WHO's recent recommendation that fresh stool is an acceptable sample for GeneXpert TB testing in children, especially small children who often cannot produce an adequate sputum sample. Diagnosing TB in this age group is difficult, and many cases are missed, leading to unacceptable rates of TB illness and death. In our large cohort of children from Dushanbe, Tajikistan, the GeneXpert stool test was positive in 69% of proven cases of TB, and there were very few false-positive tests. We also showed that this diagnostic strategy was feasible to implement in a low-middle-income country with an inefficient health care delivery system. We hope that many more programs will adopt this form of diagnosing TB in children.

Genipin Selectively Inhibits TNF-α-activated VCAM-1 But Not ICAM-1 Expression by Upregulation of PPAR-γ in Human Endothelial Cells.

Vascular inflammation process has been suggested to be an important risk factor in the development of atherosclerosis. Recently we reported that induction of peroxisome proliferator-activated receptor-γ (PPAR-γ) selectively inhibits vascular cell adhesion molecule-1 (VCAM-1) but not intercellular cell adhesion molecule-1 (ICAM-1) in tumor necrosis factor (TNF)-α-activated human umbilical vein endothelial cells (HUVEC). In this study, we investigated whether genipin inhibits expression of cellular adhesion molecules, which is relevant to inflammation. Pretreatment with genipin reduced reactive oxygen species (ROS) production and expression of VCAM-1, but not ICAM-1 in TNF-α-activated HUVEC. Genipin dose- and time-dependently increased PPAR-γ expression and inhibited TNF-α-induced phosphorylation of Akt and PKC with different degrees. Finally, genipin prevented TNF-α-induced adhesion of U937 monocytic cells to HUVEC. Taken together, these results indicate that upregualtion of PPAR-γ by genipin selectively inhibits TNF-α-induced expression of VCAM-1, in which regulation of Akt and/or PKC play a key role. We concluded that genipin can be used for the treatment of cardiovascular disorders such as atherosclerosis.

7,8-didehydrocimigenol from Cimicifugae rhizoma inhibits TNF-α-induced VCAM-1 but not ICAM-1expression through upregulation of PPAR-γ in human endothelial cells.

Activators of PPAR have been demonstrated to inhibit the induction of VCAM-1 but not ICAM-1 in human endothelial cells (EC). During the screening of anti-inflammatory activity of traditional herbs, we found 7,8-didehydrocimigenol (7,8-DHC), one of active triterpenoids of Cimicifugae rhizoma (C. rhizoma) increases PPAR-γ expression in EC in a time- and dose-dependent manner. Therefore, we asked whether 7,8-DHC selectively inhibits the expression of VCAM-1 but not ICAM-1 in TNF-α-activated EC via upregulation of PPAR-γ. Treatment with 7,8-DHC or PPAR-γ agonists (GW1929, troglitazone) inhibited the expression of VCAM-1 but not ICAM-1. Furthermore, the selective inhibition of VCAM-1 expression was inhibited by PPAR-γ antagonist, GW9662, or siPPAR-γ-transfected cells. 7,8-DHC significantly inhibited NF-kB activity via inhibition of phosphorylation of IkB and it also inhibited phosphorylation of ERK1/2 and Akt but not PKC. Finally, attachment of monocytes (U937) to EC by TNF-α was significantly reduced by 7,8-DHC. These results indicate that upregualtion of PPAR-γ by 7,8-DHC in EC inhibits NF-kB activity of TNF-α-activated EC which leads to selective inhibition of VCAM-1 expression. In addition, ERK1/2 and Akt signal pathways are involved in differential regulation by 7,8-DHC. We concluded that 7,8-DHC can be used for the treatment of cardiovascular disorders such as atherosclerosis.

Carbon monoxide from CORM-2 reduces HMGB1 release through regulation of IFN-ß/JAK2/STAT-1/INOS/NO signaling but not COX-2 in TLR-activated macrophages.

Reduction of high-mobility group box 1 (HMGB1) and NO levels may be important therapeutic strategy for treatment of sepsis. Recently, we found that carbon monoxide (CO) can reduce HMGB1 levels in septic animal models. Here, we tried to elucidate the molecular machinery of how CO inhibits HMGB1 release in toll-like receptor (TLR)-activated macrophages. Carbon monoxide-releasing molecule 2 (CORM-2) specifically inhibited the expression of iNOS (NO), but not of cyclooxygenase 2 (COX-2) (PGE2) in RAW 264.7 cells activated either by peptidoglycan (TLR-2 agonist), polyinosinic-polycytidylic acid (TLR-3 agonist), or LPS (TLR-4 agonist); this inhibition seemed to be mediated via the JAK2/STAT1 pathway. Treatment with neutralizing antibody to IFN-ß, a JAK2 inhibitor (AG490), or a STAT1 inhibitor (fludarabine) selectively inhibited iNOS, but not COX-2 in this system. Moreover, deletion of STAT1 by siRNA also showed preferential inhibition of iNOS but not COX-2 in LPS-treated cells. Carbon monoxide-releasing molecule 2 reduced IFN-ß production and phosphorylation of JAK2 and STAT1 in LPS-activated RAW264.7 cells. Carbon monoxide-releasing molecule 2 failed to inhibit iNOS and HMGB1 levels in the presence of recombinant IFN-ß and NO donor (NOC-18), respectively. Finally, plasma levels of HMGB1 and iNOS protein expression in lung tissues of cecal ligation and puncture-induced septic mice were decreased in the presence of CORM-2. Taken together, it is concluded that CO selectively inhibits iNOS over COX-2, at least through IFNß/JAK2/STAT1 signals, and this regulation plays an important role in the CORM-2-mediated inhibitory effect on HMGB1 release in macrophages.