Podocyte-specific deletion of ubiquitin carboxyl-terminal hydrolase L1 causes podocyte injury by inducing endoplasmic reticulum stress.

Ubiquitin carboxyl-terminal hydrolase L1 (UCHL1) is a unique component of the ubiquitin-proteasome system (UPS), which has multiple activities in maintaining intracellular ubiquitin levels. We previously reported the aberrant low expression of UCHL1 in podocytes of non-immune complex-mediated glomerulonephritis, and recent studies indicate that anti-UCHL1 antibody was responsible for the refractory minimal change disease (MCD), but the specific effect of UCHL1 to the podocytopathy has not been determined. Therefore, we generated podocyte-specific UCHL1 gene knockout (UCHL1cre/cre) rats model. Podocyte-specific UCHL1 knockout rats exhibited severe kidney damage, including segmental/global glomerulosclerosis, kidney function damage and severe proteinuria, compared with littermate control. Subsequently, by carrying out mass spectrometry analysis of isolated glomeruli of rats, abnormal protein accumulation of ECM-receptor Interaction was found in UCHL1cre/cre rats. Mechanistic studies in vivo and in vitro revealed that aberrant protein accumulation after UCHL1 deficiency induced endoplasmic reticulum (ER) stress, unfolded protein reaction (UPR) to reduce the protein level of podocyte skeleton proteins, and CHOP mediated apoptosis as well, which related to the dysfunction of the ubiquitin-proteasome system with decreased free monomeric ubiquitin level, thereby affecting protein ubiquitination and degradation. In addition, inhibition of ER stress by 4-PBA could attenuate the degree of ER stress and podocyte dysfunction. Our study indicates that UCHL1 is a potential target for preventing podocytes injury in some non-immune complex-mediated glomerulopathy.

Quantifying Podocyte Number in a Small Sample Size of Glomeruli with CUBIC to Evaluate Podocyte Depletion of db/db Mice.

The loss of podocyte is crucial for diagnosis and prognosis of diabetic kidney disease, whereas commonly two-dimensional methods for quantifying podocyte number existed with issues of low fidelity and accuracy. In this study, clear, unobstructed brain imaging cocktails and computational analysis (CUBIC), one of three-dimensional optical clearing approaches, was used which combines tissue clearing, immunolabeling, and a light-sheet microscope to image and evaluate podocytes in C57BL/6 (C57) and db/db mice. We discovered that 77 podocytes per glomerulus were in C57 mice. On the subject of db/db mice, there were 74 podocytes by the age of 8 w, 72 podocytes by the age of 12 w, and 66 podocytes by the age of 16 w, compared with 76 podocytes in the control group, suggesting that there was a significant decrease in podocyte number in db/db mice with the age of 16 w, showing a trend which positively correlated to the deterioration of kidney function. Sample size estimation using the PASS software revealed that taking 5%, 7.5%, and 10% of the mean podocyte number per glomerulus as the statistical allowable error and 95% as total confidence interval, 33, 15, and 9 glomeruli were independently needed to be sampled in C57 mice to represent the overall glomeruli to calculate podocyte number. Furthermore, in the control group of db/db mice, 36, 18, and 11 glomeruli were needed, compared with 46, 24, and 14 glomeruli in db/db mice by the age of 8 w, 43, 21, and 12 glomeruli by the age of 12 w, and 52, 27, and 16 by the age of 16 w. These findings indicated that precise quantification of podocyte number could judge the progression of diabetic kidney disease. In addition, a small number of glomeruli could be actually representative of the whole sample size, which indicated apparent practicability of CUBIC for clinical use.

Verteporfin inhibits the dedifferentiation of tubular epithelial cells via TGF-ß1/Smad pathway but induces podocyte loss in diabetic nephropathy.

AIMS: The dedifferentiation of tubular epithelial cells has been identified as an important trigger of renal fibrosis. The Hippo pathway is a crucial regulator of cell proliferation and differentiation. In this study, we determined the role of Hippo proteins in tubular dedifferentiation in diabetic nephropathy (DN). MAIN METHODS: In this study, we measured dedifferentiation markers and Hippo proteins in db/db mice and high glucose treated tubular epithelial cells. Then, verteporfin and knockdown of large tumor suppressor kinase (LATS) 1 and 2 were performed to uncover therapeutic targets for DN. KEY FINDINGS: Here, we found dedifferentiation and upregulated Hippo proteins in tubular epithelial cells in DN model both in vivo and in vitro. Both verteporfin and LATS knockdown could inhibit the tubular mesenchymal transition, but verteporfin showed broad inhibitory effect on Hippo proteins, especially nuclear YAP, and exacerbated podocyte loss of DN. LATS2 knockdown did not reverse the tubular E-Cadherin loss while it also induced podocyte apoptosis. Overall, intervention of LATS1 inhibited tubular dedifferentiation efficiently without affecting YAP and bringing podocyte apoptosis. Further mechanistic investigations revealed that the TGF-ß1/Smad, instead of the YAP-TEAD-CTGF signaling, might be the underlying pathway through which verteporfin and LATS1 engaged in the tubular dedifferentiation. SIGNIFICANCE: In conclusion, verteporfin is not a suitable treatment for DN owing to evitable podocyte loss and apoptosis. Targeting LATS1 is a better choice worthy of further investigation for DN therapy.

Vps34 Inhibits Hepatocellular Carcinoma Invasion by Regulating Endosome-Lysosome Trafficking via Rab7-RILP and Rab11.

PURPOSE: The role of vacuolar protein sorting 34 (Vps34), an indispensable protein required for cell vesicular trafficking, in the biological behavior of hepatocellular carcinoma (HCC) has yet to be studied. MATERIALS AND METHODS: In the present study, the expression of Vps34 in HCC and the effect of Vps34 on HCC cell invasion was detected both in vivo and in vitro. Furthermore, by modulating the RILP and Rab11, which regulate juxtanuclear lysosome aggregation and recycling endosome respectively, the underlying mechanism was investigated. RESULTS: Vps34 was significantly decreased in HCC and negatively correlated with the HCC invasiveness both in vivo and in vitro. Moreover, Vps34 could promote lysosomal juxtanuclear accumulation, reduce the invasive ability of HCC cells via the Rab7-RILP pathway. In addition, the deficiency of Vps34 in HCC cells affected the endosome-lysosome system, resulting in enhanced Rab11 mediated endocytic recycling of cell surface receptor and increased invasion of HCC cells. CONCLUSION: Our study reveals that Vps34 acts as an invasion suppressor in HCC cells, and more importantly, the endosome-lysosome trafficking regulated by Vps34 has the potential to become a target pathway in HCC treatment.

Podocyte infolding glomerulopathy with undifferentiated connective tissue disease: a case report.

Podocyte infolding glomerulopathy (PIG) is a special type of glomerular disease that has been proposed in recent years and has attracted considerable attention. PIG is characterized by the formation of microspheres and microtubules in thickened glomerular basement membrane (GBM) on electron microscopy (EM), which is recognized as podocyte cytoplasmic infolding to the GBM. However, to date, only a few cases of PIG have been reported. Herein, we report a case of a 33-year-old female with PIG with undifferentiated connective tissue disease (UCTD) in China and review the literature.