Deletion of the Ste20-like kinase SLK in skeletal muscle results in a progressive myopathy and muscle weakness

Background The Ste20-like kinase, SLK, plays an important role in cell proliferation and cytoskeletal remodeling. In fibroblasts, SLK has been shown to respond to FAK/Src signaling and regulate focal adhesion turnover through Paxillin phosphorylation. Full-length SLK has also been shown to be essential for embryonic development. In myoblasts, the overexpression of a dominant negative SLK is sufficient to block myoblast fusion. Methods In this study, we crossed the Myf5-Cre mouse model with our conditional SLK knockout model to delete SLK in skeletal muscle. A thorough analysis of skeletal muscle tissue was undertaken in order to identify defects in muscle development caused by the lack of SLK. Isometric force analysis was performed on adult knockout mice and compared to age-matched wild-type mice. Furthermore, cardiotoxin injections were performed followed by immunohistochemistry for myogenic markers to assess the efficiency muscle regeneration following SLK deletion. Results We show here that early deletion of SLK from the myogenic lineage does not markedly impair skeletal muscle development but delays the regenerative process. Interestingly, adult mice (~6 months) display an increase in the proportion of central nuclei and increased p38 activation. Furthermore, mice as young as 3 months old present with decreased force generation, suggesting that the loss of SLK impairs myofiber stability and function. Assessment of structural components revealed aberrant localization of focal adhesion proteins, such as FAK and paxillin. Our data show that the loss of SLK results in unstable myofibers resulting in a progressive myopathy. Additionally, the loss of SLK resulted in a delay in muscle regeneration following cardiotoxin injections. Conclusions Our results show that SLK is dispensable for muscle development and regeneration but is required for myofiber stability and optimal force generation

Synthesis of Novel Key Chromophoric Intermediates via C-C Coupling Reactions

The fundamentals of Pd-catalyzed Csp2−Csp2 Miyaura borylation, Suzuki cross-coupling, and Stille cross-coupling reactions for a variety of borylated precursors based on phenothiazine (PTZ), phenoxazine (POZ), carbazole (Cz), and quinoxaline (QX) units have been explored. Three palladium-based catalysts were chosen for this study: Pd(PPh3)4, Pd(PPh3)2Cl2, and Pd(dppf)Cl2, applying different reaction conditions. Around 16 desired chromophores were successfully designed and synthesized using C-C cross-coupling reactions in moderate to excellent yields, including PTZ, POZ, and Cz units coupled with QX, indolinium iodide, thienyl, phenyl, or triphenylamine moieties. Additionally, PTZ, POZ, and Cz have been employed in synthesizing various pinacol boronate ester derivatives in good to moderate yields. Interestingly, Pd(dppf)Cl2 was found to be the best catalyst for borylation, and C-C cross-coupling reactions occurred in as little as 30 min, with an excellent yield exceeding 98%. Pd(PPh3)4 and Pd(PPh3)2Cl2 catalyzed the reaction to obtain the desired products in moderate to good yields after a long time (20–24 h). On the other hand, the Suzuki-Miyaura cross-coupling between N-(2-methyl)hexyl carbazole pinacol boronate ester derivative 10c and three halogenated quinoxaline derivatives—4-(3-(5-bromothiophen-2-yl)quinoxalin-2-yl)benzaldehyde (27), 4-(5-(3-(5-bromothiophen-2-yl)quinoxalin-2-yl)thiophen-2-yl)benzaldehyde (30), and 4-(3-chloroquinoxalin-2-yl)benzaldehyde (25) catalyzed by Pd(PPh3)4—afforded three carbazole-quinoxaline chromophores (28, 30, and 31, respectively) in 2–3 h, with good to excellent yields reaching 86%. The electron-deficient QX couplers proved to be coupled efficiently using the Stille coupling reaction, which involves the coupling between electron-rich orgaostannane and electron-deficient halide. The synthesized precursors and desired chromophores were characterized by FTIR, 1H-NMR, 13C-NMR, and HRMS

Physicians’ attitudes and confidence toward dementia care: A cross-sectional study at primary healthcare facilities in the Eastern Province, Saudi Arabia

BACKGROUND: Primary care physicians play an essential role in the health of older adults as they are frequently the first point of contact. Their positive attitude and knowledge influence the quality of care provided to patients with dementia and their caregivers. This study examined the attitudes of primary care physicians towards dementia care and their confidence in their own dementia-care skills. MATERIALS AND METHODS: This cross-sectional study was conducted among 316 primary care physicians working in Eastern Province of Saudi Arabia. Data were collected using a structured questionnaire that included questions related to demographic characteristics, Dementia Care Attitude Scale (DCAS) to assess attitudes towards dementia, and Confidence in Dementia Care Skills (CDCS) Scale to measure confidence. Data were analyzed using SPSS version 29; mean and standard deviation (SD) were computed for continuous and categorical variables were described using frequencies and percentages. Mann Whitney U test and Kruskal Wallis test were used to compare attitude and confidence scores by categorical variables. RESULTS: The mean DCAS score was 36.4 ± 5.41 out of 50. On a scale ranging from 15 to 75, the mean CDCS was 51.89 ± 10.20. A statistically significant (P < 0.05) relation was found between confidence and professional rank, knowing close relatives with dementia, and number of dementia and elderly patients treated. Overall, 78.9% of physicians lacked confidence to prescribe memory medications; 32% felt that dementia management was generally more frustrating than rewarding. CONCLUSION: Primary care physicians had a positive attitude toward caring for patients with dementia. However, they lacked confidence in their dementia care skills in several areas. The confidence in their diagnostic skills was higher than their management skills. Most challenging skills were recognizing and managing behavioral symptoms of dementia. Need to develop educational and training interventions that target healthcare providers to help improving dementia care in primary care settings

Focal Adhesion Kinase Inhibitors in Combination with Erlotinib Demonstrate Enhanced Anti-Tumor Activity in Non-Small Cell Lung Cancer.

Blockade of epidermal growth factor receptor (EGFR) activity has been a primary therapeutic target for non-small cell lung cancers (NSCLC). As patients with wild-type EGFR have demonstrated only modest benefit from EGFR tyrosine kinase inhibitors (TKIs), there is a need for additional therapeutic approaches in patients with wild-type EGFR. As a key component of downstream integrin signalling and known receptor cross-talk with EGFR, we hypothesized that targeting focal adhesion kinase (FAK) activity, which has also been shown to correlate with aggressive stage in NSCLC, would lead to enhanced activity of EGFR TKIs. As such, EGFR TKI-resistant NSCLC cells (A549, H1299, H1975) were treated with the EGFR TKI erlotinib and FAK inhibitors (PF-573,228 or PF-562,271) both as single agents and in combination. We determined cell viability, apoptosis and 3-dimensional growth in vitro and assessed tumor growth in vivo. Treatment of EGFR TKI-resistant NSCLC cells with FAK inhibitor alone effectively inhibited cell viability in all cell lines tested; however, its use in combination with the EGFR TKI erlotinib was more effective at reducing cell viability than either treatment alone when tested in both 2- and 3-dimensional assays in vitro, with enhanced benefit seen in A549 cells. This increased efficacy may be due in part to the observed inhibition of Akt phosphorylation when the drugs were used in combination, where again A549 cells demonstrated the most inhibition following treatment with the drug combination. Combining erlotinib with FAK inhibitor was also potent in vivo as evidenced by reduced tumor growth in the A549 mouse xenograft model. We further ascertained that the enhanced sensitivity was irrespective of the LKB1 mutational status. In summary, we demonstrate the effectiveness of combining erlotinib and FAK inhibitors for use in known EGFR wild-type, EGFR TKI resistant cells, with the potential that a subset of cell types, which includes A549, could be particularly sensitive to this combination treatment. As such, further evaluation of this combination therapy is warranted and could prove to be an effective therapeutic approach for patients with inherent EGFR TKI-resistant NSCLC

Combination treatment with erlotinib and FAK inhibitor is more effective in reducing colony growth <i>in vitro</i> and tumor growth <i>in vivo</i>.

<p>(<b>A</b>) 3-dimensional colony formation was assessed from NSCLC cells embedded in growth factor reduced BME in 4-well chamber slides in the presence of either vehicle control (DMSO), erlotinib (10 μM), PF-228 (1 μM or 5 μM), or the combinations of erlotinib and PF-228 in complete media. Media with fresh drug was replenished every 3 days and colony size was assessed after 13 days. Images representative of two independently performed experiments are shown. (<b>B</b>) Mean colony area ± SEM is presented with statistically significant differences in mean colony size determined by unpaired Student’s <i>t</i> tests (** <i>P</i> = 0.0025, A549; * <i>P</i> = 0.044, H1299) from two independently performed experiments. (<b>C</b>) FAK inhibitors PF-228 and PF-271 show similar effectiveness in reducing cell viability <i>in vitro</i>. A549 cells were treated with either inhibitor for 48 hours and cell viability was assessed by MTT assay. Data indicates the mean ± SEM for log[inhibitor] vs. normalized response from two independently performed experiments. (<b>D</b>) <i>In vivo</i> data was collected from CD-1 nude mice injected subcutaneously with A549 cells and treated by oral gavage beginning 3 days post tumor cell injection. Data presented is the mean ± SEM for tumor volume from 4 mice per treatment each with cells injected into both hind flanks (n = 8 tumors/treatment), with statistically significant differences in tumor volume at week 8 determined by ANOVA (* P < 0.05, *** P < 0.001).</p