Neurotropin protects rotator cuff tendon cells from lidocaine-induced cell death

Background Local anesthetics often are used in rotator cuff tears as therapeutic tools, although some cases have reported that they have detrimental effects. Neurotropin (NTP) is used widely in Japan as a treatment for various chronic pain conditions and is shown to have protective effects on cartilage and nerve cells. In this study, we investigated the protective effect of NTP against lidocaine-induced cytotoxicity. Methods Tenocytes from rotator cuff tendons were incubated with lidocaine, NTP, lidocaine with NTP, and a control medium. Cell viability was evaluated using the WST-8 assay. Cell apoptosis was detected via annexin V staining using flow cytometry. The expression of BCL-2 and cytochrome c, which are involved in the intrinsic mitochondrial pathway of apoptosis, was evaluated via Western blotting and immunohistochemical staining. Results In the cell viability assay, lidocaine decreased cell viability in a dose-dependent manner, and NTP did not affect cell viability. Moreover, NTP significantly inhibited the cytotoxic effect of lidocaine. The flow cytometry analysis showed that lidocaine significantly induced apoptosis in tenocytes, and NTP considerably inhibited this lidocaine-induced apoptosis. Western blotting experiments showed that lidocaine decreased the protein expression of BCL-2, and that NTP conserved the expression of BCL-2, even when used with lidocaine. Immunohistochemical staining for cytochrome c showed that 0.1% lidocaine increased cytochrome c-positive cells, and NTP suppressed lidocaine-induced cytochrome c expression. Conclusions NTP suppresses lidocaine-induced apoptosis of tenocytes by inhibiting the mitochondrial apoptotic pathway. Intra-articular/ bursal injection of NTP with lidocaine could protect tenocytes in rotator cuff tendons against lidocaine-induced apoptosis

Implementasi Permendagri Nomor 15 Tahun 2008 Tentang Pengarusutamaan Gender pada Jenjang Pendidikan Dasar di Kota Malang

Windra Rizkiyana1 & Wahyu Widodo21 Mahasiswa & 2Staf Pengajar Program Pasca Sarjana, Universitas Muhammadiyah MalangAlamat Korespondensi : Jl. Bandung No.1 MalangEmail: [email protected] education, still found a gender gap regarding both aspects of the expansion of educationalaccess and equity, quality and relevance of education and management. The purpose of this studywere: (1) describe the substance Permendagri No. 15 of 2008 on Gender Mainstreaming; (2) describethe implementation of Permendagri No. 15 of 2008 on Gender Mainstreaming in Elementary Educationin Malang; (3) Analyze the obstacles encountered in implementation Permendagri No. 15 of 2008 onGender Mainstreaming in Elementary Education in Malang. This type of research is a descriptiveanalysis, using a qualitative approach that is supported by a quantitative approach. And the techniquesof data acolllection through by interviews and the documents. Study sites are in Malang EducationDepartment. Analysis of the data used is descriptive analysis of qualitative and quantitative theorysupported by Gender Analysis Pathway (GAP), Content Analysis and Root Analysis. Implementationof Permendagri No 15 of 2008 about gender mainstreaming in basic education levels in Malang hasnot been optimal. These proved by the remains of gender inequality or gap that occurs in all threeaspects, that access and educational equity, quality and relevance of education, as well as accountabilityand governance. Constraints encountered in implementation Permendagri No. 15 of 2008 on gendermainstreaming in elementary education in Malang include: (a) Outreach activities that are specificallyabout the PUG in primary education has not been done; (b) The budget is not specifically formainstreaming activities; (c) newly formed working group PUG.Key word: Permendagri No. 15 of 2008, gender mainstreaming, basic educatio

Monkeys mutant for PKD1 recapitulate human autosomal dominant polycystic kidney disease.

Autosomal dominant polycystic kidney disease (ADPKD) caused by PKD1 mutations is one of the most common hereditary disorders. However, the key pathological processes underlying cyst development and exacerbation in pre-symptomatic stages remain unknown, because rodent models do not recapitulate critical disease phenotypes, including disease onset in heterozygotes. Here, using CRISPR/Cas9, we generate ADPKD models with PKD1 mutations in cynomolgus monkeys. As in humans and mice, near-complete PKD1 depletion induces severe cyst formation mainly in collecting ducts. Importantly, unlike in mice, PKD1 heterozygote monkeys exhibit cyst formation perinatally in distal tubules, possibly reflecting the initial pathology in humans. Many monkeys in these models survive after cyst formation, and cysts progress with age. Furthermore, we succeed in generating selective heterozygous mutations using allele-specific targeting. We propose that our models elucidate the onset and progression of ADPKD, which will serve as a critical basis for establishing new therapeutic strategies, including drug treatments

Genetic analysis of the regulation of the voltage-gated calcium channel homolog Cch1 by the γ subunit homolog Ecm7 and cortical ER protein Scs2 in yeast

<div><p>The yeast Cch1/Mid1 Ca²⁺ channel is equivalent to animal voltage-gated Ca²⁺ channels and activated in cells incubated in low Ca²⁺ medium. We herein investigated the third subunit, Ecm7, under the same cell culture conditions. The deletion of <i>ECM7</i> slightly lowered Ca²⁺ influx activity in the <i>CNB1</i>⁺ background, in which calcineurin potentially dephosphorylates Cch1, but markedly lowered this activity in the <i>cnb1</i>Δ background. The deletion of the C-terminal cytoplasmic region of Ecm7 also reduced Ca²⁺ influx activity. We identified a novel Cch1-interacting protein, Scs2, which is known as a cortical endoplasmic reticulum membrane protein. The deletion of <i>SCS2</i> did not affect Ca²⁺ influx activity when calcineurin was inhibited by FK506, but enhanced this activity by 35% when the enzyme was not inhibited. However, this enhancement was canceled by the deletion of <i>ECM7</i>. These results suggest that Cch1/Mid1 is regulated differentially by Ecm7 and Scs2 in a manner that is dependent on the phosphorylation status of Cch1.</p></div

Monkeys mutant for PKD1 recapitulate human autosomal dominant polycystic kidney disease

ゲノム編集技術を用いてカニクイザルモデルにおいて常染色体優性多発性嚢胞腎(ADPKD)の病態再現に成功 --小動物では病態再現できない難病の研究に新たな道--. 京都大学プレスリリース. 2019-12-13.Autosomal dominant polycystic kidney disease (ADPKD) caused by PKD1 mutations is one of the most common hereditary disorders. However, the key pathological processes underlying cyst development and exacerbation in pre-symptomatic stages remain unknown, because rodent models do not recapitulate critical disease phenotypes, including disease onset in heterozygotes. Here, using CRISPR/Cas9, we generate ADPKD models with PKD1 mutations in cynomolgus monkeys. As in humans and mice, near-complete PKD1 depletion induces severe cyst formation mainly in collecting ducts. Importantly, unlike in mice, PKD1 heterozygote monkeys exhibit cyst formation perinatally in distal tubules, possibly reflecting the initial pathology in humans. Many monkeys in these models survive after cyst formation, and cysts progress with age. Furthermore, we succeed in generating selective heterozygous mutations using allele-specific targeting. We propose that our models elucidate the onset and progression of ADPKD, which will serve as a critical basis for establishing new therapeutic strategies, including drug treatments

The deletion of <i>SCS2</i> increases Ca^<i>2+</i> accumulation in calcineurin-active cells, but not in calcineurin-inactivated cells.

<p>Experimental conditions and procedures were the same as those described in the legend to <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0181436#pone.0181436.g002" target="_blank">Fig 2</a>. Cells were incubated for 2 h with 6 μM α-factor only (A) or with 6 μM α-factor and 2.0 μg/ml FK506 (B). *1, <i>p</i> < 0.05 (<i>scs2</i>Δ <i>vs</i>. WT); *2, <i>p</i> > 0.05 (<i>scs2</i>Δ <i>ecm7</i>Δ <i>vs</i>. <i>ecm7</i>Δ); *3, <i>p</i> > 0.05 (<i>scs2</i>Δ <i>vs</i>. WT); *4, <i>p</i> < 0.05 (<i>scs2</i>Δ <i>ecm7</i>Δ <i>vs</i>. <i>ecm7</i>Δ). Data are the mean ± SD of three independent experiments.</p

The deletion of <i>ECM7</i> and <i>CNB1</i> does not affect the amount of Cch1.

<p>Cells with various deletion mutations were exposed for 2 h to 6 μM α-factor and crude extracts were prepared. The amount of Cch1 in these cells was assessed by Western blotting. Enolase is an internal marker.</p

C-terminal truncations after amino acid residues 322 and 412 decrease Ecm7 activity.

<p>Experimental conditions and procedures were the same as those described in the legend to <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0181436#pone.0181436.g002" target="_blank">Fig 2</a>. The positions of a deletion (Δ429–432) and small truncation (1–432) were also described under the graph. *1, <i>p</i> > 0.05 (<i>cnb1</i>Δ<i>/</i>vector <i>vs</i>. <i>ecm7</i>Δ <i>cnb1</i>Δ<i>/</i>mutated <i>ECM7s</i>); *2, <i>p</i> < 0.05 (<i>cnb1</i>Δ<i>/</i>vector <i>vs</i>. <i>ecm7</i>Δ <i>cnb1</i>Δ<i>/</i>mutated <i>ECM7s</i>). Data are the mean ± SD of three independent experiments.</p