Targeted intracellular delivery of antitubercular bioactive(s) to Mtb infected macrophages via transferrin functionalized nanoliposomes.

The packaging of antimicrobials/chemotherapeutics into nanoliposomes can enhance their activity while minimizing toxicity. However, their use is still limited owing to inefficient/inadequate loading strategies. Several bioactive(s) which are non ionizable, and poorly aqueous soluble cannot be easily encapsulated into aqueous core of liposomes by using conventional means. Such bioactive(s) however could be encapsulated in the liposomes by forming their water soluble molecular inclusion complex with cyclodextrins. In this study, we developed Rifampicin (RIF) - 2-hydroxylpropyl-ß-cyclodextrin (HP-ß-CD) molecular inclusion complex. The HP-ß-CD-RIF complex interaction was assessed by using computational analysis (molecular modeling). The HP-ß-CD-RIF complex and Isoniazid were co-loaded in the small unilamellar vesicles (SUVs). Further, the developed system was functionalized with transferrin, a targeting moiety. Transferrin functionalized SUVs (Tf-SUVs) could preferentially deliver their payload intracellularly in the endosomal compartment of macrophages. In in vitro study on infected Raw 264.7 macrophage cells revealed that the encapsulated bioactive(s) could eradicate the pathogen more efficiently than free bioactive(s). In vivo studies further revealed that the Tf-SUVs could accumulate and maintain intracellular bioactive(s) concentrations in macrophages. The study suggests Tf-SUVs as a promising module for targeted delivery of a drug combination with improved/optimal therapeutic index and effective clinical outcomes.

HDAC6 inhibition attenuates renal injury by reducing IL-1ß secretion and RIP kinase mediated necroptosis in acute oxalate nephropathy.

BACKGROUND: Pathogenesis of acute kidney injury is driven by necro-inflammation, which is comprised of IL-1ß mediated inflammation and RIP-1 mediated tubular necroptosis. HDAC6 is reported to regulate both inflammation and cell death. In the present study, we explored the role of HDAC6 in the lysosomal exocytosis of IL-1ß and RIP-1 mediated necroptosis in the context of oxalate nephropathy. METHODS: Raw 264.7 macrophages and NRK52E stimulated with oxalate crystals and LPS with or without HDAC6 inhibitor for in vitro experiments. Acute oxalate nephropathy was induced in C57BL/6 mice by injecting sodium oxalate (75 mg/kg). For the drug intervention study, Tubastain A (TSA) was given an hour before injection of sodium oxalate. Mice were sacrificed 24 hrs after the oxalate injection, blood and kidney were harvested. Blood samples were analyzed for BUN and IL-1ß levels. Renal tissues were analyzed for histology, immunohistochemistry, RNA, and protein expression. RESULTS: HDAC6 and IL-1ß upregulated in crystal stimulated macrophages and acute oxalate nephropathy. Pre-treatment of macrophages with TSA reduced IL-1ß in supernatant without affecting the expression of pro-IL-1ß and mature IL-1ß in cell lysate. The effect of TSA on IL-1ß secretion was influenced by tubulin acetylation. Renal epithelial cell NRK52E stimulated with crystals showed upregulation of necroptosis pathway markers and concentration-dependent cell death. TSA inhibited RIP-1, RIP3, and MLKL expression along with p-MLKL in stimulated epithelial cells. TSA treatment of oxalate nephropathy mice showed decreased inflammation and tubular cell death by regulating IL-1ß and necroptosis and reduced renal injury. CONCLUSION: This study highlights the role of HDAC6 in regulating the tubulin-mediated secretion of IL-1ß and RIP kinase mediated necroptosis in acute oxalate nephropathy.

Cyclo(Val-Pro) and Cyclo(Leu-Hydroxy-Pro) from Pseudomonas sp. (ABS-36) alleviates acute and chronic renal injury under in vitro and in vivo models (Ischemic reperfusion and unilateral ureter obstruction).

The study aimed to identify small molecules having potentiality in alleviating renal injury. Two natural compounds cyclo(Val-Pro) (1) and cyclo(Leu-Hydroxy-Pro) (2) were first evaluated under acute renal injury model of ischemic reperfusion at different doses of 25, 50 and 75 mg/kg body weight. Further, the compounds were subjected to antimycin A-induced ischemic in vitro study (NRK-52E cell lines). Both the compounds significantly decreased plasma IL-1ß levels (P < 0.05). Also, the mRNA expression levels of inflammatory markers (TNF-α, IL-6 and IL-1ß) and renal injury markers (KIM-1, NGAL, α-GST and π-GST) in the renal tissues were significantly alleviated (P < 0.01) along with the improvement in histological damage and control over neutrophil infiltration as a result of ischemic reperfusion. The in vitro study revealed the protective effect against antimycin A-induced cytotoxicity (P < 0.05) and antiapoptotic effect acting through the regulation of Bax, caspase 3 (pro and cleaved) and BCL2 with reduction in Annexin+PI+ cells. Further, the compound cyclo(Val-Pro) (1) was evaluated (50 mg/kg body weight dose) in chronic unilateral ureter obstruction model of renal injury in mice and TGF-ß-induced in vitro fibrotic model (NRK-49F cell lines). Cyclo(Val-Pro) (1) significantly reduced the expression levels of fibrotic markers (collagen-1, α-SMA and TGF-ß) and showed marked alleviation of renal fibrosis (sirius red staining). Also, the proliferation of TGF-ß-induced NRK-49F cells was significantly reduced along with decreased levels of collagen-1 and α-SMA in immunohistochemistry studies. In conclusion, the compounds significantly abrogated ischemic injury by inhibiting renal inflammation and tubular epithelial apoptosis. Further, cyclo (Val-Pro) (1) exhibited significant anti-fibrotic activity through the inhibition of fibroblast activation and proliferation. Thus, these proline-based cyclic dipeptides are recommended as drug leads for treating renal injury.