Tuberous sclerosis complex exhibits a new renal cystogenic mechanism

Tuberous sclerosis complex (TSC) is a tumor predisposition syndrome with significant renal cystic and solid tumor disease. While the most common renal tumor in TSC, the angiomyolipoma, exhibits a loss of heterozygosity associated with disease, we have discovered that the renal cystic epithelium is composed of type A intercalated cells that have an intact Tsc gene that have been induced to exhibit Tsc‐mutant disease phenotype. This mechanism appears to be different than that for ADPKD. The murine models described here closely resemble the human disease and both appear to be mTORC1 inhibitor responsive. The induction signaling driving cystogenesis may be mediated by extracellular vesicle trafficking.TSC renal cystic disease develops in about half of the patients. The disease appears to caused by an induction mechanism such that a small population of mutant cells can cause significant renal cystic disease comprised of mostly genetically normal cells.Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/147796/1/phy213983.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/147796/2/phy213983_am.pd

Defective Surfactant Secretion in a Mouse Model of Hermansky-Pudlak Syndrome

Hermansky-Pudlak syndrome (HPS) in humans represents a family of disorders of lysosome-related organelle biogenesis associated with severe, progressive pulmonary disease. Human case reports and a mouse model of HPS, the pale ear/pearl mouse (ep/pe), exhibit giant lamellar bodies (GLB) in type II alveolar epithelial cells. We examined surfactant proteins and phospholipid from ep/pe mice to elucidate the process of GLB formation. The 2.8-fold enrichment of tissue phospholipids in ep/pe mice resulted from accumulation from birth through adulthood. Tissue surfactant protein (SP)-B and -C were increased in adult ep/pe mice compared with wild-type mice (WT), whereas SP-A and -D were not different. Large aggregate surfactant (LA) from adult ep/pe mice had decreased phospholipid, SP-B, and SP-C, with no differences in SP-A and -D compared with WT. Although LA from ep/pe animals exhibited an increased total protein–to–total phospholipid ratio compared with WT, surface tension was not compromised. Phospholipid secretion from isolated type II cells showed that basal and stimulated secretion from ep/pe cells were ∼ 50% of WT cells. Together, our data indicate that GLB formation is not associated with abnormal trafficking or recycling of surfactant material. Instead, impaired secretion is an important component of GLB formation in ep/pe mice

Tracking the stochastic fate of cells of the renin lineage after podocyte depletion using multicolor reporters and intravital imaging

<div><p>Podocyte depletion plays a major role in focal segmental glomerular sclerosis (FSGS). Because cells of the renin lineage (CoRL) serve as adult podocyte and parietal epithelial cell (PEC) progenitor candidates, we generated <i>Ren1cCre/R26R-ConfettiTG/WT</i> and <i>Ren1dCre/R26R-ConfettiTG/WT</i> mice to determine CoRL clonality during podocyte replacement. Four CoRL reporters (GFP, YFP, RFP, CFP) were restricted to cells in the juxtaglomerular compartment (JGC) at baseline. Following abrupt podocyte depletion in experimental FSGS, all four CoRL reporters were detected in a subset of glomeruli at day 28, where they co-expressed de novo four podocyte proteins (podocin, nephrin, WT-1 and p57) and two glomerular parietal epithelial cell (PEC) proteins (claudin-1, PAX8). To monitor the precise migration of a subset of CoRL over a 2w period following podocyte depletion, intravital multiphoton microscopy was used. Our findings demonstrate direct visual support for the migration of single CoRL from the JGC to the parietal Bowman’s capsule, early proximal tubule, mesangium and glomerular tuft. In summary, these results suggest that following podocyte depletion, multi-clonal CoRL migrate to the glomerulus and replace podocyte and PECs in experimental FSGS.</p></div

Labeled cells of renin lineage (CoRL) co-express nephrin in glomeruli of <i>Ren1cCre /R26R-ConfettiTG/WT</i> mice with experimental FSGS.

<p>The confocal images in the left column (A-C) represent nephrin staining (magenta) detected by antibody, and 4 CoRL reporters (green, red, blue, yellow) detected without antibody. The confocal images in the right column represents the same image on the left, but for ease of viewing, all 4 confetti reporter channels have been converted to green, and nephrin has been converted to red, so that co-localization can be visualized as yellow. (A, B) At baseline: All four CoRL reporter colors are restricted to the JGC (dashed white box), and nephrin staining is restricted to the glomerular tuft. (B) all four Confetti CoRL reporters (green) are seen in the JGC, with no overlap with nephrin (red). (C, D) At D14 FSGS: (C) There is a segmental decrease in nephrin staining in the right upper quadrant of the glomerular tuft. Multi-clonal CoRL are detected in glomerular tuft, but do not co-localize with nephrin. (D) The CoRL reporters in the tuft do not co-localize with nephrin staining. (E, F) At D28 FSGS: (E) Multi-clonal CoRL are detected in the glomerular tuft. (F) CoRL reporters co-localize with nephrin, creating a yellow color (arrows indicate examples).</p

Partial podocyte replacement in <i>Ren1cCre /R26R-ConfettiTG/WT</i> mice with experimental FSGS.

<p>(A-C) Double staining was performed for the podocyte marker p57 (brown, nuclear) and counterstain with Periodic acid Schiff's (pink stains matrix, blue stains nuclei) in reporter mice at baseline (A), day 14 (D14) FSGS (B) and D28 FSGS (C). (D) Podocyte number was lower at D14 compared to baseline, and partial recovered by D28. (E) Glomerulosclerosis was highest at D14 FSGS, with a significant reduction by D28. (F) The urinary albumin to creatinine ratio (ACR) was significantly higher at D14 FSGS, with a significant decrease by D28 of FSGS.</p

Multi-colored reporters of CoRL are detected in glomerular tufts of <i>Ren1cCre /R26R-ConfettiTG/WT</i> mice with FSGS.

<p>Confocal images showing four CoRL reporter colors detected without the use of antibodies–nGFP (green), cRFP (red), mCFP (blue) and cYFP (yellow). (A) All four reporters are restricted to the JGC at baseline, and are not detected in the glomerular tuft (dashed white circles). At D14 FSGS (B) and at D28 FSGS (C), all four CoRL reporter colors were detected in a subpopulation of cells in the glomerular tufts. (D) Graph showing that the percentage of glomeruli with reporter positive CoRL within the tuft was higher at D28 of FSGS and that these glomeruli contained 2–4 clones. (E) Represenative image showing all four reporters (converted to green color) and BRDU(red) do not co-localize at baseline. BRDU is present in some tubular epithelial cells as expected, but is not readily detected in JGC or the glomerular tuft. (F) BRDU staining increased at D14 of FSGS, but BRDU positive cells are not present in the JGC and glomerular tuft. (G) At D28 of FSGS there is an increase in the number of reporter labeled cells present on the glomerular tuft, however there is no overlap of reporters with BRDU.</p