Palmitoyltransferase DHHC9 and acyl protein thioesterase APT1 modulate renal fibrosis through regulating ß-catenin palmitoylation.

palmitoylation, a reversible post-translational modification, is initiated by the DHHC family of palmitoyltransferases and reversed by several acyl protein thioesterases. However, the role and mechanisms for protein palmitoylation in renal fibrosis have not been elucidated. Here we show protein palmitoylation and DHHC9 were downregulated in the fibrotic kidneys of mouse models and chronic kidney disease (CKD) patients. Ablating DHHC9 in tubular cells aggravated, while inducing DHHC9 overexpression with adeno-DHHC9 transfection or iproniazid treatment protected against kidney fibrosis in male mouse models. Mechanistically, DHHC9 palmitoylated ß-catenin, thereby promoted its ubiquitination and degradation. Additionally, acyl protein thioesterase 1 (APT1) was induced in the fibrotic kidneys, which depalmitoylated ß-catenin, increased its abundance and nuclear translocation. Ablating tubular APT1 or inhibiting APT1 with ML348 markedly protected against unilateral ureter obstruction (UUO) or ischemia/reperfusion injury (IRI)-induced kidney fibrosis in male mice. This study reveals the regulatory mechanism of protein palmitoylation in kidney fibrosis.

Mechanistic target of rapamycin complex 1 orchestrates the interplay between hepatocytes and Kupffer cells to determine the outcome of immune-mediated hepatitis.

The cell-cell interaction between hepatocytes and Kupffer cells (KCs) is crucial for maintaining liver homeostasis, and the loss of KCs and hepatocytes is known to represent a common pathogenic phenomenon in autoimmune hepatitis. Until now, the mechanisms of cell-cell interaction between hepatocytes and KCs involved in immune-mediated hepatitis remains unclear. Here we dissected the impact of activated mTORC1 on the cell-cell interaction of KCs and hepatocyte in immune-mediated hepatitis. In the liver from patients with AIH and mice administrated with Con-A, mTORC1 was activated in both KCs and hepatocytes. The activated mTORC1 signal in hepatocytes with Con-A challenge caused a markedly production of miR-329-3p. Upregulated miR-329-3p inhibited SGMS1 expression in KCs through paracrine, resulting in the death of KCs. Most of maintained KCs were p-S6 positive and distributed in hepatocyte mTORC1 negative area. The activation of mTORC1 enabled KCs expressed complement factor B (CFB) to enhance the complement alternative system, which produced more complement factors to aggravate liver injury. Our findings remonstrate a heterogeneous role of mTORC1 in specific cell type for maintaining tolerogenic liver environment, and will form the basis for the development of new interventions against immune-mediated hepatitis.

Protein phosphatase 2Acα modulates fatty acid oxidation and glycolysis to determine tubular cell fate and kidney injury.

Energy metabolism is crucial in maintaining cellular homeostasis and adapting to stimuli for tubular cells. However, the underlying mechanisms remain largely unknown. Here, we report that PP2Acα was upregulated in damaged tubular cells from patients and animal models with acute or chronic kidney injury. Using in vitro and in vivo model, we demonstrated that PP2Acα induction in damaged tubular cells suppresses fatty acid oxidation and promotes glycolysis, leading to cell death and fibrosis. Mechanistically, we revealed that PP2Acα dephosphorylates ACC through interaction with B56δ, leading to the regulation of fatty acid oxidation. Furthermore, PP2Acα also dephosphorylates p-Glut1 (Thr478) and suppresses Trim21-mediated Glut1 ubiquitination and degradation, leading to the promotion of glucose intake and glycolysis. Thus, this study adds new insight into the tubular cell metabolic alterations in kidney diseases. PP2Acα may be a promising therapeutic target for kidney injury.

Complement factor B in high glucose-induced podocyte injury and diabetic kidney disease.

The role and mechanisms for upregulating complement factor B (CFB) expression in podocyte dysfunction in diabetic kidney disease (DKD) are not fully understood. Here, analyzing Gene Expression Omnibus GSE30528 data, we identified genes enriched in mTORC1 signaling, CFB, and complement alternative pathways in podocytes from patients with DKD. In mouse models, podocyte mTOR complex 1 (mTORC1) signaling activation was induced, while blockade of mTORC1 signaling reduced CFB upregulation, alternative complement pathway activation, and podocyte injury in the glomeruli. Knocking down CFB remarkably alleviated alternative complement pathway activation and DKD in diabetic mice. In cultured podocytes, high glucose treatment activated mTORC1 signaling, stimulated STAT1 phosphorylation, and upregulated CFB expression, while blockade of mTORC1 or STAT1 signaling abolished high glucose-upregulated CFB expression. Additionally, high glucose levels downregulated protein phosphatase 2Acα (PP2Acα) expression, while PP2Acα deficiency enhanced high glucose-induced mTORC1/STAT1 activation, CFB induction, and podocyte injury. Taken together, these findings uncover a mechanism by which CFB mediates podocyte injury in DKD.

PP2Acα promotes macrophage accumulation and activation to exacerbate tubular cell death and kidney fibrosis through activating Rap1 and TNFα production.

Macrophage accumulation and activation play an essential role in kidney fibrosis; however, the underlying mechanisms remain to be explored. By analyzing the kidney tissues from patients and animal models with kidney fibrosis, we found a large induction of PP2Acα in macrophages. We then generated a mouse model with inducible macrophage ablation of PP2Acα. The knockouts developed less renal fibrosis, macrophage accumulation, or tubular cell death after unilateral ureter obstruction or ischemic reperfusion injury compared to control littermates. In cultured macrophages, PP2Acα deficiency resulted in decreased cell motility by inhibiting Rap1 activity. Moreover, co-culture of PP2Acα-/- macrophages with tubular cells resulted in less tubular cell death attributed to downregulated Stat6-mediated tumor necrosis factor α (TNFα) production in macrophages. Together, this study demonstrates that PP2Acα promotes macrophage accumulation and activation, hence accelerates tubular cell death and kidney fibrosis through regulating Rap1 activation and TNFα production.

Ovarian Index of KM Mice Influenced by Longer Term Consumption of Microwave-Heated Milk.

ABSTRACT: Microwave technology has been widely used in the food industry, but the effect of microwave-heated food on human health is being questioned. Female KM mice were chosen to be treated with microwave-heated milk (MM), and reproductive markers such as litter size, birth rate, survival rate, and ovarian index were evaluated. With longer term feeding, the reproductive status (body weight, birth rate, litter size, neonatal survival rate, interpregnancy interval, and brain superoxide dismutase and catalase activity) of KM mice treated with MM did not significantly change except for the ovarian index of first-generation mice, which was decreased significantly compared with the control group and the group given electrically heated milk. Longer term consumption of MM can affect the ovarian index of reproductive mice.

Rheb1 protects against cisplatin-induced tubular cell death and acute kidney injury via maintaining mitochondrial homeostasis.

Ras homolog enriched in brain (Rheb1), a small GTPase, plays a crucial role in regulating cell growth, differentiation, and survival. However, the role and mechanisms for Rheb1 in tubular cell survival and acute kidney injury (AKI) remain unexplored. Here we found that Rheb1 signaling was activated in kidney tubule of AKI patients and cisplatin-treated mice. A mouse model of tubule-specific deletion of Rheb1 (Tubule-Rheb1-/-) was generated. Compared to control littermates, Tubule-Rheb1-/- mice were phenotypically normal within 2 months after birth but developed more severe kidney dysfunction, tubular cell death including apoptosis, necroptosis and ferroptosis, mitochondrial defect and less PGC-1α expression after cisplatin injection. In primary cultured tubular cells, Rheb1 ablation exacerbated cisplatin-induced cell death and mitochondrial defect. Furthermore, haploinsufficiency for Tsc1 in tubular cells led to Rheb1 activation and mitigated cisplatin-induced cell death, mitochondrial defect and AKI. Together, this study uncovers that Rheb1 may protect against cisplatin-induced tubular cell death and AKI through maintaining mitochondrial homeostasis.

Fibroblast mTOR/PPARγ/HGF axis protects against tubular cell death and acute kidney injury.

Kidney fibroblasts play a crucial role in dictating tubular cell fate and the outcome of acute kidney injury (AKI). The underlying mechanisms remain to be determined. Here, we found that mTOR signaling was activated in fibroblasts from mouse kidneys with ischemia/reperfusion injury (IRI). Ablation of fibroblast Rheb or Rictor promoted, while ablation of fibroblast Tsc1 protected against tubular cell death and IRI in mice. In tubular cells cultured with conditioned media (CM) from Rheb-/- or Rictor-/- fibroblasts, less hepatocyte growth factor (HGF) receptor c-met signaling activation or staurosporine-induced cell apoptosis was observed. While CM from Tsc1-/- fibroblasts promoted tubular cell c-met signaling activation and inhibited staurosporine-induced cell apoptosis. In kidney fibroblasts, blocking mTOR signaling downregulated the expression of peroxisome proliferator-activated receptor gamma (PPARγ) and HGF. Downregulating fibroblast HGF expression or blocking tubular cell c-met signaling facilitated tubular cell apoptosis. Notably, renal PPARγ and HGF expression was less in mice with fibroblast Rheb or Rictor ablation, but more in mice with fibroblast Tsc1 ablation than their littermate controls, respectively. Together, these data suggest that mTOR signaling activation in kidney fibroblasts protects against tubular cell death and dictates the outcome of AKI through stimulating PPARγ and HGF expression.

The signaling protein Wnt5a promotes TGFß1-mediated macrophage polarization and kidney fibrosis by inducing the transcriptional regulators Yap/Taz.

M2 macrophage polarization is known to underlie kidney fibrosis. We previously reported that most of the members of the Wnt family of signaling proteins are induced in fibrotic kidneys. Dysregulation of the signaling protein Wnt5a is associated with fibrosis, but little is known about the role of Wnt5a in regulating M2 macrophage activation that results in kidney fibrosis. Here, using murine Raw 264.7 cells and bone marrow-derived macrophages, we found that Wnt5a enhanced transforming growth factor ß1 (TGFß1)-induced macrophage M2 polarization as well as expression of the transcriptional regulators Yes-associated protein (Yap)/transcriptional coactivator with PDZ-binding motif (Taz). Verteporfin blockade of Yap/Taz inhibited both Wnt5a- and TGFß1-induced macrophage M2 polarization. In mouse models of kidney fibrosis, shRNA-mediated knockdown of Wnt5a expression diminished kidney fibrosis, macrophage Yap/Taz expression, and M2 polarization. Moreover, genetic ablation of Taz in macrophages attenuated kidney fibrosis and macrophage M2 polarization in mice. Collectively, these results indicate that Wnt5a promotes kidney fibrosis by stimulating Yap/Taz-mediated macrophage M2 polarization.