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Currently, most cities situated next to rivers are facing the problem of near-water depressive spaces formation. Most frequently, coastal territories that were previously used for agricultural and industrial needs become depressive spaces. After the loss of their purpose, these facilities cease to be operated, which leads to a negative environmental impact on the environment and to its gradual degradation. The article contains the results of the study of depressive territory of the Chermyanka river section (Moscow city), in which the authors took part. The section of the river is located in the North-Eastern Administrative District of Moscow in the municipal districts of Bibirevo and Otradnoye. Based on the results of the reconnaissance survey and sociological research, the main problems of the studied depressive territory were identified. Based on the analysis of successful domestic and foreign projects on the revitalization of depressive near-water spaces, the ways of solving these problems and the development of the degraded territory are proposed. The article describes in detail the option of reorganization of the considered depressive territory, developed by the authors. The option of zoning of the river territory into green and open public spaces is justified, including an assessment of the morphometric indicators of the river, biological, sociological and landscape characteristics. The functional content of public spaces aimed at meeting the needs of various social and age groups, and communications that allow combining open public and green spaces into a holistic recreational complex are described. The new public space should perform not only a recreational function, but also a communication one. It will allow linking the Bibirevo and Otradnoye districts by a pedestrian route and will provide free movement between the banks of the river.

The few established causal genes in Alzheimer’s disease (AD), mutations in APP and PSENs, have been functionally characterized using biomarkers, capturing an in vivo profile reflecting the disease’s initial preclinical phase. SORL1, a gene encoding the endosome recycling receptor SORLA, epidemiologically behaves as a causal gene when truncating mutations lead to partial loss of protein function. Here, in an effort to test whether SORL1 can indeed function as an AD causal gene, we used CRISPR-Cas9-based gene editing to develop a novel model of SORL1 haploinsufficiency in Göttingen Minipigs taking advantage of porcine models for biomarker investigations. SORL1 haploinsufficiency in young minipigs was found to phenocopy the preclinical in vivo profile of AD observed with other causal genes, resulting in spinal fluid abnormalities in Aβ and tau, with no evident neurodegeneration or amyloid plaque formation. These studies provide functional support that SORL1 is a bona fide causal gene in AD, and when taken together with recent insight on other AD-causal genes, support the idea that dysfunctional endosomal recycling is a dominant pathogenic pathway in the disease.

Additional file 3. Figure S3: Control of RNA quality in non-AD and AD cerebella. Samples hybridization with PPIB probe showed no distinct difference in the signal from non-AD compared to AD cerebella.

Additional file 4. Figure S4: Analysis of genetic variants affecting the expression of E38b. a Representation of the boundaries of the 3′ end locus in SORL1 including localization of E38b. b eQTL mapping of SORL1 gene did not show significant association with E38b expression (SNP24: rs2282649).

The few established causal genes in Alzheimer’s disease (AD), mutations in APP and PSENs, have been functionally characterized using biomarkers, capturing an in vivo profile reflecting the disease’s initial preclinical phase. SORL1, a gene encoding the endosome recycling receptor SORLA, epidemiologically behaves as a causal gene when truncating mutations lead to partial loss of protein function. Here, in an effort to test whether SORL1 can indeed function as an AD causal gene, we used CRISPR-Cas9-based gene editing to develop a novel model of SORL1 haploinsufficiency in Gottingen Minipigs taking advantage of porcine models for biomarker investigations. SORL1 haploinsufficiency in young minipigs was found to phenocopy the preclinical in vivo profile of AD observed with other causal genes, resulting in spinal fluid abnormalities in Ab and tau, with no evident neurodegeneration or amyloid plaque formation. These studies provide functional support that SORL1 is a bona fide causal gene in AD, and when taken together with recent insight on other AD-causal genes, support the idea that dysfunctional endosomal recycling is a dominant pathogenic pathway in the disease.

Additional file 2. Figure S2: Analysis of the 3′ end of SORL1-38b. Agarose gel showing 3′ RACE products for SORL1-fl and SORL1-38b. Bands indicated with arrows have been sequenced.

Therapeutic efficacy of cisplatin-based treatment of late stage urothelial carcinoma (UC) is limited by chemoresistance. To elucidate underlying mechanisms and to develop new approaches for overcoming resistance, we generated long-term cisplatin treated (LTT) UC cell lines, characterised their cisplatin response, and determined the expression of molecules involved in cisplatin transport and detoxification, DNA repair, and apoptosis. Inhibitors of metallothioneins and Survivin were applied to investigate their ability to sensitise towards cisplatin. Cell growth, proliferation, and clonogenicity were examined after cisplatin treatment by MTT 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, EdU (5-ethynyl-2’-deoxyuridine) incorporation assay, and Giemsa staining, respectively. Cell cycle distribution and apoptosis were quantified by flow cytometry. mRNA and protein expressions were measured by real-time quantitative (qRT)-PCR, western blot, or immunofluorescence staining. LTTs recovered rapidly from cisplatin stress compared to parental cells. In LTTs, to various extents, cisplatin exporters and metallothioneins were induced, cisplatin adduct levels and DNA damage were decreased, whereas expression of DNA repair factors and specific anti-apoptotic factors was elevated. Pharmacological inhibition of Survivin, but not of metallothioneins, sensitised LTTs to cisplatin, in an additive manner. LTTs minimise cisplatin-induced DNA damage and evade apoptosis by increased expression of anti-apoptotic factors. The observed diversity among the four LTTs highlights the complexity of cisplatin resistance mechanisms even within one tumour entity, explaining heterogeneity in patient responses to chemotherapy. CA extern

Immunocytological staining with adduct-specific antibodies allows the visualization and measurement of structurally defined types of DNA damage in the nuclei of individual cells. Here we describe an immunocytological assay (ICA) procedure for the localization and quantification of such damage, in particular induced by platinum-based anticancer drugs, in cell lines , in primary cell suspensions and in frozen tissue sections.

Immunocytological staining with adduct-specific antibodies allows the visualization and measurement of structurally defined types of DNA damage in the nuclei of individual cells. Here we describe an immunocytological assay (ICA) procedure for the localization and quantification of such damage, in particular induced by platinum-based anticancer drugs, in cell lines , in primary cell suspensions and in frozen tissue sections.

Aim was to identify methylated genes with functional involvement in cisplatin-resistance development of epithelial ovarian cancer (EOC). Genome-wide analyses of hypermethylated CpG-islands in resistant cell lines in combination with qRT-PCR analyses were used to identify epigenetically silenced genes. EOC-Type-II tumors were analyzed for gene methylation and expression and TCGA data were interrogated in-silico. Experiments revealed 37 commonly hypermethylated genes in resistant cells of which Tribbles 2 (TRIB2) showed the most pronounced downregulation on mRNA level and was characterized further. TRIB2 showed a reactivation after 5'-Aza-Cytidine treatment in resistant cells but a cisplatin-dependent, prominent upregulation on mRNA level in sensitive cells, only. Re-expression in resistant A2780 cells increased the sensitivity to cisplatin and other DNA-damaging agents, but not taxanes. Contrary, knockdown of TRIB2 increased resistance to cisplatin in sensitive cells. TRIB2 was involved in the induction of a cisplatin-dependent cell cycle arrest and apoptosis by influencing p21 and survivin expression. An increased Pt-DNA-adduct formation in TRIB2 re-expressing cells did not translate in higher levels of dsDNA damage (yH2AX-foci). Thus, TRIB2 is potentially involved in the signal transduction from nucleotide excision repair of intrastrand cross links. Importantly, patient stratification of two homogenous cohorts of EOC-Type-II patients from Jena (n = 38) and the TCGA (n = 149) by TRIB2 mRNA expression consistently revealed a significantly decreased PFS for patients with low TRIB2 levels (log-rank p 0.05). Tumors from resistant patients expressed the lowest levels of TRIB2. Downregulation of TRIB2 contributes to platin-resistance and TRIB2 expression should be validated as prognostic and predictive marker for EOC.

Cisplatin is important in the treatment of various types of cancer. Although it is highly effective, it also has severe side effects, with neurotoxicity in dorsal root ganglion (DRG) neurons being one of the most common. The key mechanisms of neurotoxicity are still controversially discussed; however, disturbances of the calcium homeostasis in DRG neurons have been suggested to mediate cisplatin neurotoxicity. By using the whole-cell patch-clamp technique, immunostaining and behavioral experiments with Sprague-Dawley rats, we examined the influence of short- and long-term exposure to cisplatin on voltage-gated calcium channel (VGCC) currents (ICa(V)) in small DRG neurons. In vitro exposure to cisplatin reduced ICa(V) in a concentration-dependent manner (0.01-50μM; 13.8-77.3%; IC50 5.07μM). Subtype-specific measurements of VGCCs showed differential effects on ICa(V). While the ICa(V) of P/Q-, L- and T-type VGCCs were reduced, ICa(V) of N-type VGCCs were increased by 30.3% during depolarization to 0mV. Exposure of DRG neurons to cisplatin (0.5 or 5μM) for 24-48h in vitro significantly increased a CaMK II-mediated ICa(V) current density. Immunostaining and western blot analysis revealed an increase of N-type VGCC protein level in DRG neurons 24h after cisplatin exposure. Cisplatin-mediated activation of caspase-3 was prevented by inhibition of N-type VGCCs using Ɯ-conotoxin MVIIA. Behavioral experiments showed that Ɯ-conotoxin MVIIA treatment prevented neuropathic syndromes in vivo by inhibiting upregulation of the N-type protein level. Here we show evidence for the first time for a crucial role of N-type VGCC in the genesis of cisplatin-induced polyneuropathy.

Mutations in the amyloid-β protein precursor gene (AβPP), the presenilin 1 gene (PSEN1) or the presenilin 2 gene (PSEN2) that increase production of the AβPP-derived peptide Aβ42 cause early-onset Alzheimer's disease. Rodent models of the disease show that further increase in Aβ42 production and earlier brain pathology can be obtained by coexpressing AβPP and PSEN1 mutations. To generate such elevated Aβ42 level in a large animal model, we produced Göttingen minipigs carrying in their genome one copy of a human PSEN1 cDNA with the Met146Ile (PSEN1M146I) mutation and three copies of a human AβPP695 cDNA with the Lys670Asn/Met671Leu (AβPPsw) double-mutation. Both transgenes were expressed in fibroblasts and in the brain, and their respective proteins were processed normally. Immunohistochemical staining with Aβ42-specific antibodies detected intraneuronal accumulation of Aβ42 in brains from a 10- and an 18-month-old pig. Such accumulation may represent an early event in the pathogenesis of Alzheimer's disease.