Genetic reprogramming with stem cells regenerates glomerular epithelial podocytes in Alport syndrome.

Glomerular filtration relies on the type IV collagen (ColIV) network of the glomerular basement membrane, namely, in the triple helical molecules containing the α3, α4, and α5 chains of ColIV. Loss of function mutations in the genes encoding these chains (Col4a3, Col4a4, and Col4a5) is associated with the loss of renal function observed in Alport syndrome (AS). Precise understanding of the cellular basis for the patho-mechanism remains unknown and a specific therapy for this disease does not currently exist. Here, we generated a novel allele for the conditional deletion of Col4a3 in different glomerular cell types in mice. We found that podocytes specifically generate α3 chains in the developing glomerular basement membrane, and that its absence is sufficient to impair glomerular filtration as seen in AS. Next, we show that horizontal gene transfer, enhanced by TGFß1 and using allogenic bone marrow-derived mesenchymal stem cells and induced pluripotent stem cells, rescues Col4a3 expression and revive kidney function in Col4a3-deficient AS mice. Our proof-of-concept study supports that horizontal gene transfer such as cell fusion enables cell-based therapy in Alport syndrome.

Underestimation of glyphosate intake by the methods currently used by regulatory agencies.

The acceptable daily intake (ADI) is an estimate of the amount of a substance in food or beverages that can be consumed daily over a lifetime without presenting an appreciable risk to health. To assess the risk of ingesting glyphosate, regulatory agencies compare glyphosate daily intake to ADI. Based on published data on urine glyphosate levels measured according to known quantities of ingested glyphosate, our objectives were to test the robustness of the mathematical model currently used to calculate glyphosate daily intake, and to propose alternative models based on urinary excretion kinetics. Our results support that the quantity of ingested glyphosate is systematically underestimated by the model currently used by regulatory agencies, whereas the other models evaluated showed better estimations, with differences according to gender. Our results also show a great variability between individuals, leading to some uncertainties notably with regards to the ADI, and further support that glyphosate excretion varies significantly among individuals who follow a similar dosing regimen. In conclusion, our study highlights the lack of reliability of assessment processes carried out by regulatory agencies for glyphosate in particular, and pesticides in general, and questions the relevance of such processes supposed to safeguard human health and the environment.

Quantifiable urine glyphosate levels detected in 99% of the French population, with higher values in men, in younger people, and in farmers.

France is the first pesticide-consuming country in Europe. Glyphosate is the most used pesticide worldwide and glyphosate is detected in the general population of industrialized countries, with higher levels found in farmers and children. Little data was available concerning exposure in France. Our objective was to determine glyphosate levels in the French general population and to search for an association with seasons, biological features, lifestyle status, dietary habits, and occupational exposure. This study includes 6848 participants recruited between 2018 and 2020. Associated data include age, gender, location, employment status, and dietary information. Glyphosate was quantified by a single laboratory in first-void urine samples using ELISA. Our results support a general contamination of the French population, with glyphosate quantifiable in 99.8% of urine samples with a mean of 1.19 ng/ml + / - 0.84 after adjustment to body mass index (BMI). We confirm higher glyphosate levels in men and children. Our results support glyphosate contamination through food and water intake, as lower glyphosate levels are associated with dominant organic food intake and filtered water. Higher occupational exposure is confirmed in farmers and farmers working in wine-growing environment. Thus, our present results show a general contamination of the French population with glyphosate, and further contribute to the description of a widespread contamination in industrialized countries.

IER2-induced senescence drives melanoma invasion through osteopontin.

Expression of the immediate-early response gene IER2 has been associated with the progression of several types of cancer, but its functional role is poorly understood. We found that increased IER2 expression in human melanoma is associated with shorter overall survival, and subsequently investigated the mechanisms through which IER2 exerts this effect. In experimental melanoma models, sustained expression of IER2 induced senescence in a subset of melanoma cells in a p53/MAPK/AKT-dependent manner. The senescent cells produced a characteristic secretome that included high levels of the extracellular phosphoglycoprotein osteopontin. Nuclear localization of the IER2 protein was critical for both the induction of senescence and osteopontin secretion. Osteopontin secreted by IER2-expressing senescent cells strongly stimulated the migration and invasion of non-senescent melanoma cells. Consistently, we observed coordinate expression of IER2, p53/p21, and osteopontin in primary human melanomas and metastases, highlighting the pathophysiological relevance of IER2-mediated senescence in melanoma progression. Together, our study reveals that sustained IER2 expression drives melanoma invasion and progression through stimulating osteopontin secretion via the stochastic induction of senescence.

Spatiotemporally controlled induction of gene expression in vivo allows tracking the fate of tumor cells that traffic through the lymphatics.

Metastasis is a multistep process, during which circulating tumor cells traffic through diverse anatomical locations. Stable inducible marking of tumor cells in a manner that is tightly spatially and temporally controlled would allow tracking the contribution of cells passing through specific locations to metastatic dissemination. For example, tumor cells enter the lymphatic system and can form metastases in regional lymph nodes, but the relative contribution of tumor cells that traffic through the lymphatic system to the formation of distant metastases remains controversial. Here, we developed a novel genetic switch based on mild transient warming (TW) that allows cells to be marked in a defined spatiotemporal manner in vivo. Prior to warming, cells express only EGFP. Upon TW, the EGFP gene is excised and expression of mCherry is permanently turned on. We employed this system in an experimental pancreatic cancer model and used localized TW to induce the genetic switch in tumor cells trafficking through tumor-draining lymph nodes. Thereby we found that tumor cells disseminating via the lymphatics make a major contribution to the seeding of lung metastases. The inducible genetic marking system we have developed is a powerful tool for the tracking of metastasizing cells in vivo.

Treatment of meningioma and glioma with protons and carbon ions.

The rapid rise of particle therapy across the world necessitates evidence to justify its ever-increasing utilization. This narrative review summarizes the current status of these technologies on treatment of both meningiomas and gliomas, the most common benign and malignant primary brain tumors, respectively. Proton beam therapy (PBT) for meningiomas displays high rates of long-term local control, low rates of symptomatic deterioration, along with the potential for safe dose-escalation in select (but not necessarily routine) cases. PBT is also associated with low adverse events and maintenance of functional outcomes, which have implications for quality of life and cost-effectiveness measures going forward. Data on carbon ion radiation therapy (CIRT) are limited; existing series describe virtually no high-grade toxicities and high local control. Regarding the few available data on low-grade gliomas, PBT provides opportunities to dose-escalate while affording no increase of severe toxicities, along with maintaining appropriate quality of life. Although dose-escalation for low-grade disease has been less frequently performed than for glioblastoma, PBT and CIRT continue to be utilized for the latter, and also have potential for safer re-irradiation of high-grade gliomas. For both neoplasms, the impact of superior dosimetric profiles with endpoints such as neurocognitive decline and neurologic funcionality, are also discussed to the extent of requiring more data to support the utility of particle therapy. Caveats to these data are also described, such as the largely retrospective nature of the available studies, patient selection, and heterogeneity in patient population as well as treatment (including mixed photon/particle treatment). Nevertheless, multiple prospective trials (which may partially attenuate those concerns) are also discussed. In light of the low quantity and quality of available data, major questions remain regarding economic concerns as well.

Cell cycle quiescence can suppress transcription from an ecdysone receptor-based inducible promoter in mammalian cells.

Inducible gene expression is a powerful tool for basic research, gene therapy and biotechnology, whose utility depends in part on consistent levels of induction regardless of metabolic status or physiological context. Here we examined the inducibility of the ecdysone receptor-based RheoSwitch mammalian inducible expression system in proliferating cells and in cell cycle-arrested cells. We found that both contact inhibition and growth arrest subsequent to serum deprivation dramatically reduced the levels of induction of reporter genes that could be achieved in 3T3 fibroblasts but in not NMuMG mammary epithelial cells. These data have implications for the use of the RheoSwitch system in inducible gene expression applications.

Nsl1p is essential for the establishment of bipolarity and the localization of the Dam-Duo complex.

We identified a physical complex consisting of Mtw1p, an established kinetochore protein, with Nnf1p, Nsl1p and Dsn1p and have demonstrated that Nnf1p, Nsl1p and Dsn1p localize to the Saccharomyces cerevisiae kinetochore. When challenged prior to metaphase, the temperature-sensitive mutants nsl1-16 and nsl1-42 as well as Nsl1p-depleted cells failed to establish a bipolar spindle-kinetochore interaction and executed monopolar segregation of sister chromatids. In contrast, an nsl1-16 defect could not be evoked after the establishment of bipolarity. The observed phenotype is characteristic of that of mutants with defects in the protein kinase Ipl1p or components of the Dam-Duo kinetochore complex. However nsl1 mutants did not exhibit a defect in microtubule-kinetochore untethering as the ipl1-321 mutant does. Instead, they exhibited a severe defect in the kinetochore localization of the Dam-Duo complex suggesting this to be the cause for the failure of nsl1 cells to establish bipolarity. Moreover the analysis of Nsl1p-depleted cells indicated that Nsl1p is required for the spindle checkpoint and kinetochore integrity.