PPARα-targeted mitochondrial bioenergetics mediate repair of intestinal barriers at the host-microbe intersection during SIV infection.

Chronic gut inflammatory diseases are associated with disruption of intestinal epithelial barriers and impaired mucosal immunity. HIV-1 (HIV) causes depletion of mucosal CD4+ T cells early in infection and disruption of gut epithelium, resulting in chronic inflammation and immunodeficiency. Although antiretroviral therapy (ART) is effective in suppressing viral replication, it is incapable of restoring the "leaky gut," which poses an impediment for HIV cure efforts. Strategies are needed for rapid repair of the epithelium to protect intestinal microenvironments and immunity in inflamed gut. Using an in vivo nonhuman primate intestinal loop model of HIV/AIDS, we identified the pathogenic mechanism underlying sustained disruption of gut epithelium and explored rapid repair of gut epithelium at the intersection of microbial metabolism. Molecular, immunological, and metabolomic analyses revealed marked loss of peroxisomal proliferator-activated receptor-α (PPARα) signaling, predominant impairment of mitochondrial function, and epithelial disruption both in vivo and in vitro. To elucidate pathways regulating intestinal epithelial integrity, we introduced probiotic Lactobacillus plantarum into Simian immunodeficiency virus (SIV)-inflamed intestinal lumen. Rapid recovery of the epithelium occurred within 5 h of L. plantarum administration, independent of mucosal CD4+ T cell recovery, and in the absence of ART. This intestinal barrier repair was driven by L. plantarum-induced PPARα activation and restoration of mitochondrial structure and fatty acid ß-oxidation. Our data highlight the critical role of PPARα at the intersection between microbial metabolism and epithelial repair in virally inflamed gut and as a potential mitochondrial target for restoring gut barriers in other infectious or gut inflammatory diseases.

Disruption of mitochondrial function as mechanism for anti-cancer activity of a novel mitochondriotropic menadione derivative.

Menadione, also known as vitamin K3, is a 2-methyl-1,4 naphthoquinone with a potent cytotoxic activity mainly resulting from its quinone redox-cycling with production of reactive oxygen species (ROS). Although increased ROS generation is considered a relevant mechanism in cancer cell death, it may not be sufficiently effective to kill cancer cells due to phenotypic adaptations. Therefore, combining ROS-generating agents with other molecules targeting important cancer cell phenotypes can be an effective therapeutic strategy. As mitochondrial dysfunction has been implicated in many human diseases, including cancer, we describe here the discovery of a mitochondrial-directed agent (MitoK3), which was developed by conjugating a TPP cation to the C3 position of the menadione's naphthoquinone ring, increasing its selective accumulation in mitochondria, as well as led to alterations of its redox properties and consequent biological outcome. MitoK3 disturbed the mitochondrial bioenergetic apparatus, with subsequent loss of mitochondrial ATP production. The combinatory strategy of MitoK3 with anticancer agent doxorubicin (DOX) resulted in a degree of cytotoxicity higher than those of the individual molecules, as the combination triggered tumour apoptotic cell death evident by caspase 3/9 activities, probably through mitochondrial destabilization or by interference with mitochondrial redox processes. The results of this investigation support the importance of drug discovery process in developing molecules that can be use as adjuvant therapy in patients with specific cancer subtypes.

Identification of small molecules that improve ATP synthesis defects conferred by Leber's hereditary optic neuropathy mutations.

Inherited mitochondrial complex I mutations cause blinding Leber's hereditary optic neuropathy (LHON), for which no curative therapy exists. A specific biochemical consequence of LHON mutations in the presence of trace rotenone was observed: deficient complex I-dependent ATP synthesis (CIDAS) and mitochondrial O2 consumption, proportional to the clinical severity of the three primary LHON mutations. We optimized a high-throughput assay of CIDAS to screen 1600 drugs to 2, papaverine and zolpidem, which protected CIDAS in LHON cells concentration-dependently. TSPO and cAMP were investigated as protective mechanisms, but a conclusive mechanism remains to be elucidated; next steps include testing in animal models.

Toxins in botanical dietary supplements: blue cohosh components disrupt cellular respiration and mitochondrial membrane potential.

Certain botanical dietary supplements have been associated with idiosyncratic organ-specific toxicity. Similar toxicological events, caused by drug-induced mitochondrial dysfunction, have forced the withdrawal or U.S. FDA "black box" warnings of major pharmaceuticals. To assess the potential mitochondrial liability of botanical dietary supplements, extracts from 352 authenticated plant samples used in traditional Chinese, Ayurvedic, and Western herbal medicine were evaluated for the ability to disrupt cellular respiration. Blue cohosh (Caulophyllum thalictroides) methanol extract exhibited mitochondriotoxic activity. Used by some U.S. midwives to help induce labor, blue cohosh has been associated with perinatal stroke, acute myocardial infarction, congestive heart failure, multiple organ injury, and neonatal shock. The potential link between mitochondrial disruption and idiosyncratic herbal intoxication prompted further examination. The C. thalictroides methanol extract and three saponins, cauloside A (1), saponin PE (2), and cauloside C (3), exhibited concentration- and time-dependent mitochondriotoxic activities. Upon treatment, cell respiration rate rapidly increased and then dramatically decreased within minutes. Mechanistic studies revealed that C. thalictroides constituents impair mitochondrial function by disrupting membrane integrity. These studies provide a potential etiological link between this mitochondria-sensitive form of cytotoxicity and idiosyncratic organ damage.

Comparative study of chromatographic medium-associated mass and potential antitumor activity loss with bioactive extracts.

Natural product drug discovery programs often rely on the use of silica (Si) gel, reversed-phase media, or size-exclusion resins (e.g., RP-C18, Sephadex LH-20) for compound purification. The synthetic polymer-based sorbent Diaion HP20SS (cross-linked polystyrene matrix) is used as an alternative to prepare purified natural product libraries. To evaluate the impact of chromatographic media on the isolation of biologically active, yet chromatographically unstable natural products, Diaion HP20SS was evaluated side-by-side with normal-phase sorbents for irreversible binding of extract constituents and their effects on bioactivity. An array of chemically diverse natural product-rich extracts was selected as a test panel, and a cell-based reporter assay for hypoxia-inducible factor-1 (HIF-1) was employed to monitor potential change(s) in bioactivity. Silica gel caused significant irreversible binding of three out of 10 extracts. Curcuma longa, Saururus cernuus, and Citrus reticulata extracts showed decreased HIF-1 inhibitory activity after elution through Si gel. An additional nonpolar column wash of HP20SS with EtOAc retained considerable bioactivities of active extracts. In general, Si gel produced the greatest loss of bioactivity. However, HP20SS elution reduced significantly HIF-1 inhibitory activity of certain extracts (e.g., Asimina triloba).