Podocyte injury at young age causes premature senescence and worsens glomerular aging.

As life expectancy continues to rise, age-related diseases are becoming more prevalent. For example, proteinuric glomerular diseases typified by podocyte injury have worse outcomes in the elderly compared with young patients. However, the reasons are not well understood.

Inhibiting NLRP3 signaling in aging podocytes improves their life- and health-span.

The decrease in the podocyte's lifespan and health-span that typify healthy kidney aging cause a decrease in their normal structure, physiology and function. The ability to halt and even reverse these changes becomes clinically relevant when disease is superimposed on an aged kidney. RNA-sequencing of podocytes from middle-aged mice showed an inflammatory phenotype with increases in the NLRP3 inflammasome, signaling for IL2/Stat5, IL6 and TNF, interferon gamma response, allograft rejection and complement, consistent with inflammaging.

Upregulated PD-1 signaling antagonizes glomerular health in aged kidneys and disease.

With an aging population, kidney health becomes an important medical and socioeconomic factor. Kidney aging mechanisms are not well understood. We previously showed that podocytes isolated from aged mice exhibit increased expression of programmed cell death protein 1 (PD-1) surface receptor and its 2 ligands (PD-L1 and PD-L2).

Application of Cryopreserved Human Intestinal Mucosa and Cryopreserved Human Enterocytes in the Evaluation of Herb-Drug Interactions: Evaluation of CYP3A Inhibitory Potential of Grapefruit Juice and Commercial Formulations of Twenty-Nine Herbal Supplements.

Commercial formulations of 29 commonly used herbal supplements (HSs) and grapefruit juice were evaluated for drug interaction potential via quantification of their CYP3A inhibitory potential in two in vitro experimental models of human small intestine, cryopreserved human intestinal mucosa (CHIM), and cryopreserved human enterocytes (CHEs). Two CYP3A substrates were used-in the studies with CHIM, CYP3A activity was quantified via liquid chromatography tandem mass spectrometry quantification of midazolam 1'-hydroxylation, whereas in CHE, luciferin-IPA metabolism to luciferin was quantified by luminescence. Upon treatment of CHIM with the estimated lumen concentration of the HS upon each oral administration (manufacturers' recommended dosage dissolved in 200 ml of culture medium), >80% CYP3A inhibition was observed for green tea extract, St.

Cryopreserved Human Intestinal Mucosal Epithelium: A Novel In Vitro Experimental System for the Evaluation of Enteric Drug Metabolism, Cytochrome P450 Induction, and Enterotoxicity.

We report here a novel in vitro enteric experimental system, cryopreserved human intestinal mucosa (CHIM), for the evaluation of enteric drug metabolism, drug-drug interaction, drug toxicity, and pharmacology. CHIM was isolated from the small intestines of four human donors. The small intestines were first dissected into the duodenum, jejunum, and ileum, followed by collagenase digestion of the intestinal lumen.

A Novel In Vitro Experimental System for the Evaluation of Drug Metabolism: Cofactor-Supplemented Permeabilized Cryopreserved Human Hepatocytes (MetMax Cryopreserved Human Hepatocytes).

We report here a novel experimental system, cryopreserved MetMax human hepatocytes (MMHHs), for in vitro drug metabolism studies. MMHHs consist of cofactor-supplemented permeabilized cryopreserved human hepatocytes. The use procedures for MMHHs are significantly simplified from that for conventional cryopreserved human hepatocytes (CCHHs): 1) storage at -80°C instead of in liquid nitrogen and 2) usage directly after thawing without centrifugation and microscopic evaluation of cell density and viability and cell density adjustment.

In vitro evaluation of hepatotoxic drugs in human hepatocytes from multiple donors: Identification of P450 activity as a potential risk factor for drug-induced liver injuries.

A possible risk factor for drug-induced hepatotoxicity is drug metabolizing enzyme activity, which is known to vary among individuals due to genetic (genetic polymorphism) and environmental factors (environmental pollutants, foods, and medications that are inhibitors or inducers of drug metabolizing enzymes). We hypothesize that hepatic cytochrome P450-dependent monooxygenase (CYP) activity is one of the key risk factors for drug induced liver injuries (DILI) in the human population, especially for drugs that are metabolically activated to cytotoxic/reactive metabolites. Human hepatocytes from 19 donors were evaluated for the activities of 8 major P450 isoforms: CYP1A2, CYP2B6, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1 and CYP3A4.

Inducible costimulator (ICOS) up-regulation on activated T cells in chronic graft-versus-host disease after dog leukocyte antigen-nonidentical hematopoietic cell transplantation: a potential therapeutic target.

Background: Inducible costimulator (ICOS), a member of the CD28 family of costimulatory molecules, is induced on CD4 and CD8 T cells after their activation. ICOS functions as an essential immune regulator and ICOS blockade is a potential approach to immune modulation in allogeneic transplantation. Here, we describe the expression profile of ICOS in dogs and determine whether ICOS expression is up-regulated during chronic graft-versus-host disease (GVHD) and host-versus-graft reactions in the canine hematopoietic cell transplantation model.

Activation of Notch signaling during ex vivo expansion maintains donor muscle cell engraftment.

Transplantation of myogenic stem cells possesses great potential for long-term repair of dystrophic muscle. However, a single donor muscle biopsy is unlikely to provide enough cells to effectively transplant the muscle mass of a patient affected by muscular dystrophy. Expansion of cells ex vivo using traditional culture techniques significantly reduces engraftment potential.

Inhibition of CD26/DPP-IV enhances donor muscle cell engraftment and stimulates sustained donor cell proliferation.

Background: Transplantation of myogenic stem cells possesses great potential for long-term repair of dystrophic muscle. In murine-to-murine transplantation experiments, CXCR4 expression marks a population of adult murine satellite cells with robust engraftment potential in mdx mice, and CXCR4-positive murine muscle-derived SP cells home more effectively to dystrophic muscle after intra-arterial delivery in mdx5cv mice. Together, these data suggest that CXCR4 plays an important role in donor cell engraftment.

Antagonistic and agonistic anti-canine CD28 monoclonal antibodies: tools for allogeneic transplantation.

Background: It has been presumed that antibody-mediated selective costimulatory molecule blockade of CD28 is superior to cytotoxic T lymphocyte antigen 4 (CTLA4)-Ig. This is based on the premise that specifically blocking CD28 allows inhibitory signals through CTLA-4 to proceed, which furthermore suppresses T-cell function.

Methods: The extracelluar domain of canine (ca)CD28 was cloned from dog peripheral blood mononuclear cells.

Establishment of long-term tolerance to SRBC in dogs by recombinant canine CTLA4-Ig.

Background: Blockade of the CD28 costimulatory molecule by recombinant human cytotoxic T lymphocyte (CTL)-associated antigen (CTLA4)-Ig or CD40-CD154 interaction with the monoclonal antibody 5C8 together with donor-specific transfusion led to enhanced engraftment in the canine model of dog leukocyte antigen (DLA)-identical marrow transplantation after 1 Gy total body irradiation. To reduce or eliminate total body irradiation conditioning regimens, we have sought to develop canine specific reagents.

Methods: We have created a fusion protein of the extracellular domain of canine (c) CTLA-4 linked to the hinge-CH2-CH3 domains of canine IgG1 in a pcDNA3.

Long-term tolerance to kidney allografts in a preclinical canine model.

Durable immune tolerance supporting vascularized allotransplantation offers the possibility of extending graft survival and avoiding harmful complications of chronic immunosuppression. Immune tolerance to renal allografts was induced in a preclinical canine model through engraftment of donor hematopoietic cells using a combination of low-dose total body irradiation and a short course of immunosuppression. Subsequently, donor renal allografts were transplanted accompanied by bilateral native nephrectomies.

Molecular cloning and characterization of canine ICOS.

Inducible costimulatory receptor (ICOS) is one recently identified member of the CD28 family of costimulatory molecules. Evidence suggests ICOS functions as a critical immune regulator and, to evaluate these effects, we employed the canine model system that has been used to develop strategies currently in clinical use for hematopoietic stem cell transplantation. To investigate the effects of blocking the ICOS pathway in the canine hematopoietic cell transplantation model, we tested existing murine and human reagents and cloned the full length of the open reading frame of canine ICOS cDNA to allow the development of reagents specific for the canine ICOS.