Rapamycin attenuates PLA2R activation-mediated podocyte apoptosis via the PI3K/AKT/mTOR pathway

Membranous nephropathy (MN) is the most common cause of nephrotic syndrome in adults without diabetes. Primary MN has been associated with circulating antibodies against native podocyte antigens, including phospholipase A2 receptor (PLA2R); however, precision therapy targeting the signaling cascade of PLA2R activation is lacking. Both PLA2R and the mammalian target of rapamycin (mTOR) exist in podocytes, but the interplay between these two proteins and their roles in MN warrants further exploration. This study aimed to investigate the crosstalk between PLA2R activation and mTOR signaling in a human podocyte cell line. We demonstrated that podocyte apoptosis was induced by Group IB secretory phospholipase A2 (sPLA2IB) in a concentration- and time-dependent manner via upregulation of phosphoinositide 3-kinase (PI3K), protein kinase B (AKT), and mTOR, and inhibited by rapamycin or LY294002. Furthermore, aberrant activation of the PI3K/AKT/mTOR pathway triggers both extrinsic (caspase-8 and caspase-3) and intrinsic (Bcl-2-associated X protein [BAX], B-cell lymphoma 2 [BCL-2], cytochrome c, caspase-9, and caspase-3) apoptotic cascades in podocytes. The therapeutic implications of our findings are that strategies to reduce PLA2R activation and PI3K/AKT/mTOR pathway inhibition in PLA2R-activated podocytes help protect podocytes from apoptosis. The therapeutic potential of rapamycin shown in this study provides cellular evidence supporting the repurposing of rapamycin for MN treatment

Extracardiac septum transversum/proepicardial endothelial cells pattern embryonic coronary arterio–venous connections

Recent reports suggest that mammalian embryonic coronary endothelium (CoE) originates from the sinus venosus and ventricular endocardium. However, the contribution of extracardiac cells to CoE is thought to be minor and nonsignificant for coronary formation. Using classic (Wt1(Cre)) and previously undescribed (G2-Gata4(Cre)) transgenic mouse models for the study of coronary vascular development, we show that extracardiac septum transversum/proepicardium (ST/PE)-derived endothelial cells are required for the formation of ventricular coronary arterio-venous vascular connections. Our results indicate that at least 20% of embryonic coronary arterial and capillary endothelial cells derive from the ST/PE compartment. Moreover, we show that conditional deletion of the ST/PE lineage-specific Wilms' tumor suppressor gene (Wt1) in the ST/PE of G2-Gata4(Cre) mice and in the endothelium of Tie2(Cre) mice disrupts embryonic coronary transmural patterning, leading to embryonic death. Taken together, our results demonstrate that ST/PE-derived endothelial cells contribute significantly to and are required for proper coronary vascular morphogenesi

Deletion of Wt1 during early gonadogenesis leads to differences of sex development in male and female adult mice

Assessing the role of the WT1 transcription factor (WT1) during early gonad differentiation and its impact on adult sex development has been difficult due to the complete gonadal agenesis and embryonic lethality exhibited by Wt1KO mouse models. Here, we generated Wt1LoxP/GFP;Wt1Cre mice, the first Wt1KO mouse model that reaches adulthood with a dramatically reduced Wt1 expression during early gonadogenesis. Wt1LoxP/GFP;Wt1Cre mice lacked mature gonads and displayed genital tracts containing both male and female genital structures and ambiguous genitalia. We found that WT1 is necessary for the activation of both male and female sex-determining pathways, as embryonic mutant gonads failed to upregulate the expression of the genes specific for each genetic programme. The gonads of Wt1LoxP/GFP;Wt1Cre mice showed a lack of production of Sertoli and pre-granulosa cells and a reduced number of germ cells. NR5A1 and the steroidogenic genes expression was modulated differently in XY and XX Wt1LoxP/GFP;Wt1Cre gonads, explaining the mutant phenotypes. Further studies of the XX Wt1LoxP/GFP;Wt1Cre gonads revealed that deletion of WT1 at an early stage impaired the differentiation of several cell types including somatic cells and the ovarian epithelium. Through the characterisation of this Wt1KO mouse model, we show that the deletion of Wt1 during early gonadogenesis produces dramatic defects in adult sex development

In vivo imaging of the tumor and its associated microenvironment using combined CARS / 2-photon microscopy

The use of confocal and multi-photon microscopy for intra-vital cancer imaging has impacted on our understanding of cancer cell behavior and interaction with the surrounding tumor microenvironment in vivo. However, many studies to-date rely on the use fluorescent dyes or genetically encoded probes that enable visualization of a structure or cell population of interest, but do not illuminate the complexity of the surrounding tumor microenvironment. Here, we show that multi-modal microscopy combining 2-photon fluorescence with CARS can begin to address this deficit, enabling detailed imaging of the tumor niche without the need for additional labeling. This can be performed on live tumor-bearing animals through optical observation windows, permitting real-time and longitudinal imaging of dynamic processes within the tumor niche

Empagliflozin Protects HK-2 Cells from High Glucose-Mediated Injuries via a Mitochondrial Mechanism

Empagliflozin is known to retard the progression of kidney disease in diabetic patients. However, the underlying mechanism is incompletely understood. High glucose induces oxidative stress in renal tubules, eventually leading to mitochondrial damage. Here, we investigated whether empagliflozin exhibits protective functions in renal tubules via a mitochondrial mechanism. We used human proximal tubular cell (PTC) line HK-2 and employed western blotting, terminal deoxynucleotidyl transferase dUTP nick end labelling assay, fluorescence staining, flow cytometry, and enzyme-linked immunosorbent assay to investigate the impact of high glucose and empagliflozin on cellular apoptosis, mitochondrial morphology, and functions including mitochondrial membrane potential (MMP), reactive oxygen species (ROS) production, and adenosine triphosphate (ATP) generation. We found that PTCs were susceptible to high glucose-induced mitochondrial fragmentation, and empagliflozin ameliorated this effect via the regulation of mitochondrial fission (FIS1 and DRP1) and fusion (MFN1 and MFN2) proteins. Empagliflozin reduced the high glucose-induced cellular apoptosis and improved mitochondrial functions by restoring mitochondrial ROS production, MMP, and ATP generation. Our results suggest that empagliflozin may protect renal PTCs from high glucose-mediated injuries through a mitochondrial mechanism. This could be one of the novel mechanisms explaining the benefits demonstrated in EMPA-REG OUTCOME trial

Visceral and subcutaneous fat have different origins and evidence supports a mesothelial source

International audience: Fuelled by the obesity epidemic, there is considerable interest in the developmental origins of white adipose tissue (WAT) and the stem and progenitor cells from which it arises. Whereas increased visceral fat mass is associated with metabolic dysfunction, increased subcutaneous WAT is protective. There are six visceral fat depots: perirenal, gonadal, epicardial, retroperitoneal, omental and mesenteric, and it is a subject of much debate whether these have a common developmental origin and whether this differs from that for subcutaneous WAT. Here we show that all six visceral WAT depots receive a significant contribution from cells expressing Wt1 late in gestation. Conversely, no subcutaneous WAT or brown adipose tissue arises from Wt1-expressing cells. Postnatally, a subset of visceral WAT continues to arise from Wt1-expressing cells, consistent with the finding that Wt1 marks a proportion of cell populations enriched in WAT progenitors. We show that all visceral fat depots have a mesothelial layer like the visceral organs with which they are associated, and provide several lines of evidence that Wt1-expressing mesothelium can produce adipocytes. These results reveal a major ontogenetic difference between visceral and subcutaneous WAT, and pinpoint the lateral plate mesoderm as a major source of visceral WAT. They also support the notion that visceral WAT progenitors are heterogeneous, and suggest that mesothelium is a source of adipocytes

Demographic and Clinical Characteristics of Patients with Severe Asthma in the Asian Pacific Region : data from the International Severe Asthma Registry (ISAR)

Peer reviewe

Acute Multiple Organ Failure in Adult Mice Deleted for the Developmental Regulator Wt1

There is much interest in the mechanisms that regulate adult tissue homeostasis and their relationship to processes governing foetal development. Mice deleted for the Wilms' tumour gene, Wt1, lack kidneys, gonads, and spleen and die at mid-gestation due to defective coronary vasculature. Wt1 is vital for maintaining the mesenchymal–epithelial balance in these tissues and is required for the epithelial-to-mesenchyme transition (EMT) that generates coronary vascular progenitors. Although Wt1 is only expressed in rare cell populations in adults including glomerular podocytes, 1% of bone marrow cells, and mesothelium, we hypothesised that this might be important for homeostasis of adult tissues; hence, we deleted the gene ubiquitously in young and adult mice. Within just a few days, the mice suffered glomerulosclerosis, atrophy of the exocrine pancreas and spleen, severe reduction in bone and fat, and failure of erythropoiesis. FACS and culture experiments showed that Wt1 has an intrinsic role in both haematopoietic and mesenchymal stem cell lineages and suggest that defects within these contribute to the phenotypes we observe. We propose that glomerulosclerosis arises in part through down regulation of nephrin, a known Wt1 target gene. Protein profiling in mutant serum showed that there was no systemic inflammatory or nutritional response in the mutant mice. However, there was a dramatic reduction in circulating IGF-1 levels, which is likely to contribute to the bone and fat phenotypes. The reduction of IGF-1 did not result from a decrease in circulating GH, and there is no apparent pathology of the pituitary and adrenal glands. These findings 1) suggest that Wt1 is a major regulator of the homeostasis of some adult tissues, through both local and systemic actions; 2) highlight the differences between foetal and adult tissue regulation; 3) point to the importance of adult mesenchyme in tissue turnover