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Nearest-neighbour clustering is a simple yet powerful machine learning algorithm that finds natural application in the decoding of signals in classical optical-fibre communication systems. Quantum k-means clustering promises a speed-up over the classical k-means algorithm; however, it has been shown to not currently provide this speed-up for decoding optical-fibre signals due to the embedding of classical data, which introduces inaccuracies and slowdowns. Although still not achieving an exponential speed-up for NISQ implementations, this work proposes the generalised inverse stereographic projection as an improved embedding into the Bloch sphere for quantum distance estimation in k-nearest-neighbour clustering, which allows us to get closer to the classical performance. We also use the generalised inverse stereographic projection to develop an analogous classical clustering algorithm and benchmark its accuracy, runtime and convergence for decoding real-world experimental optical-fibre communication data. This proposed ‘quantum-inspired’ algorithm provides an improvement in both the accuracy and convergence rate with respect to the k-means algorithm. Hence, this work presents two main contributions. Firstly, we propose the general inverse stereographic projection into the Bloch sphere as a better embedding for quantum machine learning algorithms; here, we use the problem of clustering quadrature amplitude modulated optical-fibre signals as an example. Secondly, as a purely classical contribution inspired by the first contribution, we propose and benchmark the use of the general inverse stereographic projection and spherical centroid for clustering optical-fibre signals, showing that optimizing the radius yields a consistent improvement in accuracy and convergence rate.

Patellar tendinopathy (jumper's knee) has a high prevalence in jumping athletes. Excessive load on the patellar tendon through high volumes of training and competition is an important risk factor. Structural changes in the tendon are related to a higher risk of developing patellar tendinopathy. The critical tendon load that affects tendon structure is unknown. The aim of this study was to investigate patellar tendon structure on each day of a 5-day volleyball tournament in an adolescent population (16-18 years). The right patellar tendon of 41 players in the Australian Volleyball Schools Cup was scanned with ultrasound tissue characterization ( UTC) on every day of the tournament ( Monday to Friday). UTC can quantify structure of a tendon into four echo types based on the stability of the echo pattern. Generalized estimating equations ( GEE) were used to test for change of echo type I and II over the tournament days. Participants played between eight and nine matches during the tournament. GEE analysis showed no significant change of echo type percentages of echo type I ( Wald chi-square = 4.603, d.f. = 4, P = 0.331) and echo type II ( Wald chi-square = 6.070, d.f. = 4, P = 0.194) over time. This study shows that patellar tendon structure of 16-18-year-old volleyball players is not affected during 5 days of cumulative loading during a volleyball tournament. [ABSTRACT FROM AUTHOR]

Objectives: To describe a post platelet-rich plasma (PRP) injection, exercise-based physical therapy program, investigate feasibility and report the first results of patellar tendinopathy patients treated with PRP injection combined with the physical therapy program. Study Design: Case-series. Setting: A PRP injection followed by a physical therapy program seems promising for the treatment of patellar tendinopathy. However, descriptions of physical therapy programs are often limited and incomplete. Participants: Five patellar tendinopathy patients (six tendons) in the degenerative phase. Main outcome measure: VISA-P score. Results: Muscle strength, endurance, power and retraining sport-specific function form the basis for the physical therapy program aiming to improve the load capacity of the knee. The program is characterised by gradually increasing intensity and difficulty of exercises. Five of the six tendons showed an improvement of at least 30 points on the VISA-P after 26 weeks. Conclusions: This study extensively describes, based on current knowledge, a physical therapy program after PRP injection for patellar tendinopathy patients. The combination treatment reported in this study is feasible and seems to be promising for patients in the late/degenerative phase of patellar tendinopathy. [ABSTRACT FROM AUTHOR]

Objective Injection treatments are increasingly used as treatment for patellar tendinopathy. The aim of this systematic review is to describe the different injection treatments, their rationales and the effectiveness of treating patellar tendinopathy. Methods A computerised search of the Medline, Embase, CINAHL and Web of Knowledge databases was conducted on 1 May 2010 to identify studies on injection treatments for patellar tendinopathy. Results 11 articles on seven different injection treatments (dry needling, autologous blood, high-volume, platelet-rich plasma, sclerosis, steroids and aprotinin injections) were found: 4 randomised controlled trials (RCTs), 1 non-RCT, 4 prospective cohort studies and 2 retrospective cohort studies. All studies reported positive results. The Delphi scores of the four RCTs ranged from 5 to 8 out of 9. Different and sometimes contradictory rationales were used for the injection treatments. Conclusion All seven different injection treatments seem promising for treating patellar tendinopathy. Unlike the other injection treatments, steroid treatment often shows a relapse of symptoms in the long term. Results should be interpreted with caution as the number of studies is low, few high-quality studies have been conducted and the studies are hard to compare due to different methodology. More high-quality studies using the same cross-cultural reliable and valid outcome measure are needed, as well as further research into the pathophysiology. Finally, some implications are provided for clinicians who want to use injection treatments as a part of their treatment for patellar tendinopathy, distinguishing between reactive and degenerative phase of patellar tendinopathy. [ABSTRACT FROM AUTHOR]

Patellar tendinopathy (PT) is an injury with a high prevalence in sports. Knowledge of risk factors is essential for developing preventive measures and rehabilitation programmes. However, risk factors associated with PT have not yet been systematically studied. This review was undertaken to identify risk factors associated with PT. The literature was systematically searched to identify articles that investigated risk factors for PT. There was no strong or moderate evidence that any investigated risk factor was associated with PT. For nine risk factors there was some evidence: weight, body mass index, waist-to-hip ratio, leg-length difference, arch height of the foot, quadriceps flexibility, hamstring flexibility, quadriceps strength and vertical jump performance. Based on the present evidence, reducing body weight, increasing upper-leg flexibility and quadriceps strength and the use of orthotics may be beneficial treatment options. However, it should be stressed that the evidence for the nine identified risk factors was only limited. Therefore, there is a clear need for high-quality studies in order to identify the exact risk factors associated with PT. [ABSTRACT FROM AUTHOR]

Background Patellar tendinopathy (PT) is an injury which has a high prevalence in sports, especially in those involving jumping. Because successful treatment of PT remains challenging, prevention is of utmost importance. Before effective prevention programs can be developed, it is important to identify the risk factors that play a role in the development of this injury. Objective The aim of this study was to review the literature and to identify risk factors associated with PT. Design Systematic review of the literature. Setting A search of the Pubmed, EMBASE and AMED databases was conducted to identify studies. Studies were included in the systematic review if they contained: (1) empirical research that investigated factors associated with PT; and (2) a patient group with PT and a comparison group (or tested for differences across groups in a cross-sectional survey). Assessment of risk factors The methodological quality of the studies was assessed with a methodological quality assessment list. A strength of evidence criterion was used to identify risk factors. This criterion took into account the percentage of studies that found an association for a factor and the methodological quality of these studies. Results 11 studies were included in the review investigating over 40 potential risk factors. The overall methodological quality score of the studies was low. Potential risk factors were categorised in four groups; demographics, anthropometrics, sports related factors and strength/flexibility. The present review found evidence for two risk factors for PT: waist-to-hip ratio and hip and knee flexibility. Conclusion Waist-to-hip ratio and hip and knee flexibility were identified as risk factors for PT. There is a clear need for high quality studies in order to identify the exact risk factors associated with PT. [ABSTRACT FROM PUBLISHER]

Background Patellar tendinopathy has a multifactorial aetiology, and many potential risk factors have been described in the literature. Findings are conflicting though, and most research is conducted on elite athletes. Objective Aim of the study is to determine risk factors for patellar tendinopathy in a large representative sample of basketball and volleyball players. Design Cross-sectional survey using an online questionnaire. Setting Basketball and volleyball players of all playing levels. Participants All basketball and volleyball players between ages 18 and 35 years (n=7740) from the Dutch Basketball Association and the Dutch Volleyball Association were invited to complete the questionnaire. 1561 volleyball players completed the questionnaire. Assessment of risk factors The following independent variables were collected: Gender, age, body mass index, waist-to-hip ratio, years playing basketball/volleyball, hours of training during the last week, number of games during the last month, average hours of other sports per week, training increase compared to last year, playing surface, playing level and playing position. Main outcome measurements Presence of symptomatic patellar tendinopathy was the dependent variable. Results Risk factors for patellar tendinopathy were age, playing at the national level, being male and playing volleyball (compared to playing basketball). Risk factors for men and women were comparable. Among volleyball players outside hitters and middle blockers/hitters had an increased risk compared to setters. For basketball players no risk factors could be identified. No differences in risk factors were found between athletes with unilateral and bilateral patellar tendinopathy. Conclusion Several potential risk factors for patellar tendinopathy were identified. These findings should be taken into account for prevention and rehabilitation purposes. [ABSTRACT FROM PUBLISHER]

To develop a core outcome set for Achilles tendinopathy (COS-AT) for use in clinical trials we performed a five-step process including (1) a systematic review of available outcome measurement instruments, (2) an online survey on truth and feasibility of the available measurement instruments, (3) an assessment of the methodological quality of the selected outcome measurement instruments, (4) an online survey on the outcome measurement instruments as COS and (5) a consensus in-person meeting. Both surveys were completed by healthcare professionals and patients. The Outcome Measures in Rheumatology guidelines with a 70% threshold for consensus were followed. We identified 233 different outcome measurement instruments from 307 included studies; 177 were mapped within the International Scientific Tendinopathy Symposium Consensus core domains. 31 participants (12 patients) completed the first online survey (response rate 94%). 22/177 (12%) outcome measurement instruments were deemed truthful and feasible and their measurement properties were evaluated. 29 participants (12 patients) completed the second online survey (response rate 88%) and three outcome measurement instruments were endorsed: the Victorian Institute of Sports Assessment-Achilles questionnaire, the single-leg heel rise test and evaluating pain after activity using a Visual Analogue Scale (VAS, 0-10). 12 participants (1 patient) attended the final consensus meeting, and 1 additional outcome measurement instrument was endorsed: evaluating pain during activity/loading using a VAS (0-10). It is recommended that the identified COS-AT will be used in future clinical trials evaluating the effectiveness of an intervention. This will facilitate comparing outcomes of intervention strategies, data pooling and further progression of knowledge about AT. As COS-AT is implemented, further evidence on measurement properties of included measures and new outcome measurement instruments should lead to its review and refinement., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2024. No commercial re-use. See rights and permissions. Published by BMJ.)

Purpose: The purpose of this study was to develop a multidisciplinary guideline for patellofemoral pain (PFP) and patellar tendinopathy (PT) to facilitate clinical decision-making in primary and secondary care., Methods: A multidisciplinary expert panel identified questions in clinical decision-making. Based on a systematic literature search, the strength of the scientific evidence was determined according to the Grading of Recommendations, Assessment, Development and Evaluations (GRADE) method and the weight assigned to the considerations by the expert panel together determined the strength of the recommendations., Results: After confirming PFP or PT as a clinical diagnosis, patients should start with exercise therapy. Additional conservative treatments are indicated only when exercise therapy does not result in clinically relevant changes after six (PFP) or 12 (PT) weeks. Pain medications should be reserved for cases of severe pain. The additional value of imaging assessments for PT is limited. Open surgery is reserved for very specific cases of nonresponders to exercise therapy and those requiring additional conservative treatments. Although the certainty of evidence regarding exercise therapy for PFP and PT had to be downgraded ('very low GRADE' and 'low GRADE'), the expert panel advocates its use as the primary treatment strategy. The panel further formulated weaker recommendations regarding additional conservative treatments, pain medications, imaging assessments and open surgery ('very low GRADE' to 'low GRADE' assessment or absence of scientific evidence)., Conclusion: This guideline recommends starting with exercise therapy for PFP and PT. The recommendations facilitate clinical decision-making, and thereby optimizing treatment and preventing unnecessary burdens, risks and costs to patients and society., Level of Evidence: Level V, clinical practice guideline., (© 2024 The Author(s). Knee Surgery, Sports Traumatology, Arthroscopy published by John Wiley & Sons Ltd on behalf of European Society of Sports Traumatology, Knee Surgery and Arthroscopy.)

Cellular senescence is the state of permanent growth arrest. Chemotherapeutic drugs induce senescence, known as therapy-induced senescence. Although there are studies deciphering processes in senescence, more studies providing detailed information on therapy-induced senescence at the transcriptome level are needed. In order to understand temporal molecular changes of doxorubicin treatment in the course of senescence formation, two data sets from HeLa cells at 16 h and 72 h doxorubicin treatment were analyzed. GO BP enrichment, KEGG pathways and hub genes specific to or shared between 16 h and 72 h doxorubicin treated HeLa cells were identified. Genes functioning in p53 signaling were upregulated only in 16 h, while genes functioning in extracellular matrix organization were upregulated only in 72 h doxorubicin treated HeLa cells. Wound healing genes were gradually upregulated from 16 h to 72 h doxorubicin treatment and metabolic pathways were downregulated at both. ncRNA processing and ribosome biogenesis GO BP terms were enriched in upregulated genes at 16 h, while these terms were enriched in downregulated genes at 72 h senescent HeLa cells. According to our results, genes functioning in p53 signaling may be involved in the induction of senescence, but may not be required to maintain senescence in HeLa cells., Competing Interests: Declaration of competing interest The authors declare that they have no conflict of interest., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Background: Propylene chlorohydrins (PCHs) are formed as byproducts in the reaction between starch and propylene oxide (PO). For hydroxypropylated (HP) starch applications in food, Joint FAO/WHO Expert Committee on Food additives (JECFA) set the maximum allowed total propylene chlorohydrin (PHC-t) residues level at 1 mg/kg., Objective: To develop an improved analytical method for the determination of the PCH-t content in starches in the low mg/kg range to replace the outdated JECFA method., Methods: A new GC-MS method that utilizes aqueous methanol as extraction medium for PCH. The GC-MS system is equipped with a programmable temperature vaporization injector and Stabilwax-DA column and uses helium as carrier gas. Quantitative detection is achieved in selected ion monitoring mode., Results: This single-laboratory validation (SLV) study showed good linear calibrations for both 1-chloro-2-propanol (PCH-1) and 2-chloro-1-propanol (PCH-2) in the concentration range of 0.5-4 mg/kg in dry starch. The lower LOQ of PCH-1 and PCH-2 was 0.2-0.3 mg/kg in dry starch, the reproducibility RSD (RSDR) at the concentration level of 1-2 mg/kg in dry starch was 3-5%, and the recovery values for both PCH-1 and PCH-2 were in the range of 78-112% at a concentration level of about 0.6 mg/kg in dry starch. Compared with the current, outdated JECFA method, the new GC-MS method is more sustainable, less laborious, and therefore more economical. The analytical capacity of the new method is 4-5 times higher than the analytical capacity of the old JECFA method., Conclusion: The GC-MS method is fit for a multi-laboratory trial (MLT)., Highlights: Based on the results of this SLV and the MLT (will be published in a second paper), the Joint FAO/WHO Expert Committee on Food Additives has recently decided to replace the outdated GC-FID JECFA method with the new GC-MS method for the determination of PCH-t content in starches., (© The Author(s) 2023. Published by Oxford University Press on behalf of AOAC INTERNATIONAL. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Cellular senescence was first described as a state characterized by telomere shortening, resulting in limiting cell proliferation in aging. Apart from this type of senescence, which is called replicative senescence, other senescence types occur after exposure to different stress factors. One of these types of senescence induced after adjuvant therapy (chemotherapy and radiotherapy) is called therapy-induced senescence. The treatment with chemotherapeutics induces cellular senescence in normal and cancer cells in the tumor microenvironment. Thus therapy-induced senescence in the cancer microenvironment is accepted one of the drivers of tumor progression. Recent studies have revealed that senescence-associated secretory phenotype induction has roles in pathological processes such as inducing epithelial-mesenchymal transition and promoting tumor vascularization. Thus senolytic drugs that specifically kill senescent cells and senomorphic drugs that inhibit the secretory activity of senescent cells are seen as a new approach in cancer treatment. Developing and discovering new senotherapeutic agents targeting senescent cells is also gaining importance. In this review, we attempt to summarize the signaling pathways regarding the metabolism, cell morphology, and organelles of the senescent cell. Furthermore, we also reviewed the effects of SASP in the cancer microenvironment and the senotherapeutics that have the potential to be used as adjuvant therapy in cancer treatment., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Background: Nine core domains for tendinopathy have been identified. For Achilles tendinopathy there is large variation in outcome measures used, and how these fit into the core domains has not been investigated., Objective: To identify all available outcome measures outcome measures used to assess the clinical phenotype of Achilles tendinopathy in prospective studies and to map the outcomes measures into predefined health-related core domains., Design: Systematic review., Data Sources: Embase, MEDLINE (Ovid), Web of Science, CINAHL, The Cochrane Library, SPORTDiscus and Google Scholar., Eligibility Criteria for Selecting Studies: Clinical diagnosis of Achilles tendinopathy, sample size ≥ ten participants, age ≥ 16 years, and the study design was a randomized or non-randomized clinical trial, observational cohort, single-arm intervention, or case series., Results: 9376 studies were initially screened and 307 studies were finally included, totaling 13,248 participants. There were 233 (177 core domain) different outcome measures identified across all domains. For each core domain outcome measures were identified, with a range between 8 and 35 unique outcome measures utilized for each domain. The proportion of studies that included outcomes for predefined core domains ranged from 4% for the psychological factors domain to 72% for the disability domain., Conclusion: 233 unique outcome measures for Achilles tendinopathy were identified. Most frequently, outcome measures were used within the disability domain. Outcome measures assessing psychological factors were scarcely used. The next step in developing a core outcome set for Achilles tendinopathy is to engage patients, clinicians and researchers to reach consensus on key outcomes measures., Prospero Registration: CRD42020156763., (© 2021. The Author(s), under exclusive licence to Springer Nature Switzerland AG.)

Objective: The aim of this study was to evaluate the effect of finishing and polishing procedures of compomer and bulk-fill composite resins on cytotoxicity against human gingival fibroblasts by xCELLigence analysis., Study Design: Filtek™ Bulk Fill composite and Dyract XP compomer were used. After curing, the specimens were randomly divided into two groups and finishing-polishing procedures were applied to one group; no finishing-polishing procedures were applied to the other group. For the first time in this study, pure gold samples were prepared with the same weight and base area as the test specimens and the wells containing the pure gold samples were determined as the control group. xCELLigence system was used to assess the response of the human gingival fibroblasts after exposure to test specimens. Measurements were recorded for 72 hours after adding specimens., Results: Finishing and polishing procedures caused a significant increase in cell viability of Dyract XP compomer samples at all time periods; the percentage of cell viability reached above 70% after finishing and polishing procedures. However, significant effects were not observed in Filtek™ Bulk Fill composite samples at any time period., Conclusion: Finishing and polishing procedures play an essential role in increasing the biocompatibility of Dyract XP compomer. It is recommended to apply finishing and polishing procedures even though a smooth surface may be obtained in restorations with matrix strips.

Chemotherapy-induced senescent cancer cells secrete several factors in their microenvironment called SASP. Accumulated evidence states that SASP is responsible for some of the harmful effects of chemotherapy such as drug resistance and the induction of cancer cell proliferation, migration, and invasion. Therefore, to develop senolytic and/or senomorphic drugs, targeting the senescent cells gains importance as a new strategy for preventing the damage that senescent cancer cells cause. In the current work, we evaluated whether Rho/Rho kinase pathway has the potential to be used as a target pathway for the development of senolytic and/or senomorphic drugs in doxorubicin-induced senescent cancer cell lines. We have determined that inhibition of Rho/Rho kinase pathway with CT04 and Y27632 reduced the secretory activity of senescent cancer cells and changed the composition of SASP. Our results indicate that ROCK 2 isoform was responsible for these observed effects on the SASP. In addition, non-senescent cancer cell proliferation and migration accelerated by senescent cells were set back to the pre-induction levels after ROCK inhibition. Moreover, contrary to the previous observations, another important finding of the current work is that senescent HeLa and A549 cells did not engulf the non-senescent HeLa, A549 cells, and non-cancer HUVEC. These results indicate that ROCK inhibitors, in particular ROCK 2 specific inhibitors, have the potential to be developed as novel senomorphic drugs. In addition, we found that all senescent cancer cells do not share the same engulfment ability, and this process should not be generalized., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Objectives: In cancer treatment, it is important to prevent or slow down metastasis as well as preventing the proliferation of cancer cells. In this study, we aimed to find pyrazole compounds with antimigratory properties., Methods: The 'PASSonline' programme was used to determine the possible pharmacological activities of the pyrazole compounds selected from the library, and two pyrazole derivatives were identified as a transcription factor STAT inhibitor with a high probability. There are studies known that JAK/STAT pathway is related to cancer cell migration, thus the possible antimigratory effects of these two synthesized pyrazole compounds were examined in A549 cancer cells., Key Findings: Our data demonstrated that compound-2 at different concentrations significantly inhibited cell migration in A549 cells. Then, the effects of these compounds on STAT activation were evaluated. We reported that 10 µM compound-2 induced a significant phosphorylation of STAT1 suggesting that STAT1 activation may be responsible for the antimigratory effect of compound-2., Conclusions: Taken together, the compound-2 is a promising compound with the antimigratory activity for cancer treatment, and further studies are needed to synthesize more active derivatives by evaluating the structure-activity relationship of leading compound-2., (© The Author(s) 2021. Published by Oxford University Press on behalf of the Royal Pharmaceutical Society. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

During apoptosis, myosin light chain phosphorylation induced by ROCK 1, activated by caspase 3-mediated cleavage, results in the formation of membrane blebs. Additionally, actin-myosin-based contraction induced by the activation of ROCK is involved in the apoptotic nuclear disintegration. In previous studies, it was reported that ROCK 1 was only cleaved by caspase 3 in cell death and caspase 7 was involved in truncation of ROCK 1 in in-vitro cell-free conditions. Here we reported that caspase 2 is involved in the truncation of ROCK 1 directly as well as caspase 3 and caspase 7. Utilizing caspase 3-deficient MCF-7, MDA-MB-231 and HeLa cells, we demonstrated that caspase 2 produced an active fragment of approximately 130 kDa of ROCK 1 in cell death. The cleaved active fragment of ROCK 1 is also responsible for the formation of membrane blebbing in cell death. Interestingly, caspase 2-mediated cleavage of ROCK 1 might occur in the region where caspase 3 truncates ROCK 1. Moreover, the presence of an active cleaved form of ROCK 1 in the nuclei implies that this fragment might play a role in the disruption of nuclear integrity. Taken together, it was determined that caspase 2 has a role in the truncation of ROCK 1 in cell death, and a new activation mechanism has been defined for ROCK 1., Competing Interests: Declaration of competing interest The authors state that there is no conflict of interest., (Copyright © 2021 Elsevier Inc. All rights reserved.)

In drug discovery, most small molecules cannot cross many stages, only a few can become drug candidates. Once the drug molecule is approved and marketed, nontarget effects that are not easily distinguishable from the actual target of the drugs might be evaluated. This situation restricts the treatment. Thus, the discovery of new drugs is a very long and expensive process. In recent years, without developing new drugs, the approach of using different and new target molecules in new indications apart from the indications of licensed drug molecules has gained importance.In this study, using the connectivity map program, it was determined that metformin and tolbutamide used in the treatment of type II diabetes had the potential to inhibit Rho kinase. In the experimental results to confirm this data, it has been shown that metformin and tolbutamide decrease the cell area within 24 h and metformin inhibits the activation of Rho kinase in MCF-7 cells.These results indicate that metformin, which is used in the treatment of type II diabetes, acts as a ROCK inhibitor. Metformin has potential in the treatment of various pathological conditions in which Rho kinase has a role., Competing Interests: CONFLICT OF INTEREST: The authors declare that they have no conflicts of interest., (Copyright © 2021 The Author(s).)

Context: Several studies have been conducted to better understand the effect of load on the Achilles tendon structure. However, the effect of a high cumulative load consisting of repetitive cyclic movements, such as those that occur during the running of a marathon, on Achilles tendon structure is not yet clear. Clinicians, coaches, and athletes will benefit from knowledge about the effects of a marathon on the structure of the Achilles tendon., Objective: To investigate the short-term response of the Achilles tendon structure to running a marathon., Design: Case series (prospective)., Setting: Sports medicine centers., Patients or Other Participants: Ten male nonelite runners who ran in a marathon., Main Outcomes Measure(s): Tendon structure was assessed before and 2 and 7 days after a marathon using ultrasound tissue characterization (UTC), an imaging tool that quantifies tendon organization in 4 echo types (I-IV). Echo type I represents the most stable echo pattern, and echo type IV, the least stable., Results: At 7 days postmarathon, both the insertional and midportion structure changed significantly. At both sites, the percentage of echo type II increased (insertion P < .01; midportion P = .02) and the percentages of echo types III and IV decreased (type III: insertion P = .01; midportion P = .02; type IV: insertion P = .01; midportion P < .01). Additionally, at the insertion, the percentage of echo type I decreased ( P < .01)., Conclusions: We observed the effects of running a marathon on the Achilles tendon structure 7 days after the event. Running the marathon combined with the activity performed shortly thereafter might have caused the changes in tendon structure. This result emphasizes the importance of sufficient recovery time after running a marathon to prevent overuse injuries.

Purpose: Ultrasound tissue characterization (UTC) is used in research and clinical practice to quantify tendon structure of the patellar tendon. This is the first study to investigate the inter- and intra-rater reliability for UTC of the patellar tendon on a large scale., Method: Fifty participants (25 patellar tendinopathy, 25 asymptomatic) were recruited. The affected patellar tendons in symptomatic and right tendons in asymptomatic participants were scanned with UTC twice by one researcher and once by another. The same was done for contour marking (needed to analyze a UTC scan) of the tendon. Intraclass correlation coefficient (ICC (2,1)) for echo-types I, II, III, IV, aligned fibrillar structure (echo-types I + II), and disorganized structure (echo-types III + IV) were calculated. This was done for UTC scans as well as solely marking contours., Results: Inter-rater reliability showed fair to good ICC values for echo-types I (0.65) and II (0.46) and excellent ICC values for echo-type III (0.81), echo-type IV (0.83), aligned fibrillar structure (0.82), and disorganized structure (0.82). Intra-rater reliability showed excellent ICC values for echo-types I (0.76), III (0.88), IV (0.85), aligned fibrillar structure (0.88), and disorganized fibrillar structure (0.88) and a fair to good value for echo-type II (0.61). Contour marking showed excellent ICC values for all echo-types., Conclusion: This study showed that UTC scans for patellar tendons have overall good intra-rater and inter-rater reliability. To optimize reliability of UTC scans of the patellar tendon, using the same rater and using aligned fibrillar structure (echo-types I + II combined) and disorganized structure (echo-types III + IV combined) as outcome measures can be considered., (© 2019 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Recently drug-induced senescence has gained momentum as a new approach in cancer therapy. It is accepted that senescent cells display typical phenotypic features including flattened, enlarged, and multinucleated cell morphology. However, it is not well elucidated how these morphological alterations occur. The current study evaluates the possible role of Rho/Rho kinase pathway in cardiac glycoside-induced senescent cell morphology in HeLa cells. Our results indicate that the administration of cardiac glycosides, ouabain, digoxin, bufalin, to HeLa cells induced cellular senescence leading to an increase in the volume, area and maximum thickness of the cells. Although preincubation of specific Rho kinase inhibitor Y-27632 did not inhibit the occurrence of cardiac glycoside-induced senescence in cells, it reduced the cell area and cell volume. Inhibition of Rho by CT04 produced similar results as seen for the preincubation of Y-27632. In addition, inhibition of Rock caused a decrease in increased actin stress fibers in senescent cells induced by ouabain. Additionally, preincubation of Y-27632 decreased the ouabain-induced the phosphorylation of MYPT and cofilin. In conclusion, Rock inhibition-mediated alteration of senescent cell morphology may be associated with the decreased actin stress fibers formation. Since it is known that secretory activity is accompanied by the changes of cell morphology, these morphological alterations observed by the inhibition of Rho/Rho kinase pathway may also lead to important secretory functions of senescent cells., (© 2018 Wiley Periodicals, Inc.)

Objectives: The aim of this study was to investigate the effects of a 4-wk in-season exercise program of isometric or isotonic exercises on tendon structure and dimensions as quantified by ultrasound tissue characterization (UTC)., Design: This was a randomized clinical trial. Volleyball and basketball players (16-31 yrs, n = 29) with clinically diagnosed patellar tendinopathy were randomized to a 4-wk isometric or isotonic exercise program. The programs were designed to decrease patellar tendon pain. A baseline and 4-wk UTC scan was used to evaluate change in tendon structure., Results: No significant change in tendon structure or dimensions on UTC was detected after the exercise program despite patellar tendinopathy symptoms improving. The percentage and mean cross-sectional area of aligned fibrillar structure (echo types I + II) (Z = -0.414, P = 0.679) as well as disorganized structure (echo types III + IV) (Z = -0.370, P = 0.711) did not change over the 4-wk exercise program. Change in tendon structure and dimensions on UTC did not differ significantly between the groups., Conclusion: Structural properties and dimensions of the patellar tendon on UTC did not change after a 4-wk isometric or isotonic exercise program for athletes with patellar tendinopathy in-season, despite an improvement in symptoms. It seems that structural improvements are not required for a positive clinical outcome.

The aim of this study is to formulate and compare the physicochemical properties of negatively charged liposomes and poly(lactide-co-glycolide) (PLGA) nanoparticles loaded with gemcitabine hydrochloride. The influence of the formulation variables on the liposome and nanoparticle properties on particle size, zeta potential, encapsulation efficiency, and drug release was evaluated. Although the PEGylated nanoparticles and PEGylated liposomes were of the same size (∼200 nm), the encapsulation efficiency was 1.4 times higher for PEGylated liposomes than for PEGylated nanoparticles. The optimized formulation of PEGylated liposomes and PEGylated nanoparticles had 26.1 ± 0.18 and 18.8 ± 1.52% encapsulation efficiency, respectively. The release of drug from the PEGylated liposomes and PEGylated nanoparticles exhibited a biphasic pattern that was characterized by a fast initial release during the first 2 h followed by a slower continuous release. Transmission electron microscopy (TEM) images identified separate circular structures of the liposomes and nanoparticles. The in vitro cytotoxicity of the optimized formulations was assessed in MCF-7 and MDA-MB-231 cells, and the results showed that the cytotoxicity effect of the gemcitabine hydrochloride-loaded liposomes and nanoparticles was more than commercial product Gemko ® and gemcitabine hydrochloride solution.

In this study, cytotoxic effects of the dichloromethane, ethyl acetate, ethanol, and aqueous extracts of the aerial parts of Cyclotrichium niveum (Boiss.) Manden. & Scheng (Lamiaceae) were evaluated. We tested HeLa, MCF-7 cancer cells, and MRC-5 and MCF-10A normal cells. The ethyl acetate and dichloromethane extracts induced cytotoxicity whereas the ethanol and aqueous extracts had no cytotoxic activity against both cancer cells. IC 50 values of the dichloromethane extract were 353.0 ± 84.30 μg/ml, 114.8 ± 40.34 μg/ml, 39 ± 0.56 μg/ml, and 49 ± 13 μg/ml in HeLa, MCF-7, MRC-5, MCF-10A cells, respectively. IC 50 values of the ethyl acetate extract were 117.0 ± 36.24 μg/ml in HeLa cells, 156.3 ± 19.86 μg/ml in MCF-7 cells, 1100 ± 340 μg/ml in MRC-5 cells and 7900 ± 1200 μg/ml in MCF-10A cells. Additionally, the ethyl acetate extract showed more selectivity to HeLa and MCF-7 cancer cells than MRC-5 and MCF-10A normal cells. Our results of HPLC analysis showed that apigenin in the ethyl acetate extract (2.2518 ± 0.0005 mg/100 mg extract) might be responsible of that selective cytotoxic effect. In the current work, we have shown for the first time that C. niveum has cytotoxic properties in the cancer cell lines tested., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Objective: This study aimed to compare the immediate analgesic effects of 2 resistance programs in in-season athletes with patellar tendinopathy (PT). Resistance training is noninvasive, a principle stimulus for corticospinal and neuromuscular adaptation, and may be analgesic., Design: Within-season randomized clinical trial. Data analysis was conducted blinded to group., Setting: Subelite volleyball and basketball competitions., Participants: Twenty jumping athletes aged more than 16 years, participating in games/trainings 3 times per week with clinically diagnosed PT., Interventions: Two quadriceps resistance protocols were compared; (1) isometric leg extension holds at 60 degrees knee flexion (80% of their maximal voluntary isometric contraction) or (2) isotonic leg extension (at 80% of their 8 repetition maximum) 4 times per week for 4 weeks. Time under load and rest between sets was matched between groups., Main Outcome Measures: (1) Pain (0-10 numerical rating score) during single leg decline squat (SLDS), measured preintervention and postintervention sessions. (2) VISA-P, a questionnaire about tendon pain and function, completed at baseline and after 4 weeks., Results: Twenty athletes with PT (18 men, mean 22.5 ± 4.7 years) participated (isotonic n = 10, isometric n = 10). Baseline median SLDS pain was 5/10 for both groups (isotonic range 1-8, isometric range 2-8). Isometric contractions produced significantly greater immediate analgesia (P < 0.002). Week one analgesic response positively correlated with improvements in VISA-P at 4 weeks (r = 0.64)., Conclusions: Both protocols appear efficacious for in-season athletes to reduce pain, however, isometric contractions demonstrated significantly greater immediate analgesia throughout the 4-week trial. Greater analgesia may increase the ability to load or perform.

Objectives: Many athletes with patellar tendinopathy participate in sports with symptoms during or after activities. Current treatments do not decrease pain in-season; eccentric exercises in-season result in an increase in pain. This study examined if isometric and isotonic exercises relieved pain in competing athletes with patellar tendinopathy., Design: Randomised clinical trial., Methods: Jumping athletes with patellar tendinopathy playing at least three times per week participated in this study. Athletes were randomised into an isometric or isotonic exercise group. The exercise programs consisted of four isometric or isotonic exercise sessions per week for four weeks. Pain during a single leg decline squat (SLDS) on a Numeric Rating Scale (NRS; 0-10) was used as the main outcome measure; measurements were completed at baseline and at 4-week follow-up., Results: Twenty-nine athletes were included in this study. Median pain scores improved significantly over the 4-week intervention period in both the isometric group (Z=-2.527, p=0.012, r=-0.63) and isotonic group (Z=-2.952, p=0.003, r=-0.63). There was no significant difference in NRS pain score change (U=29.0, p=0.208, r=0.29) between the isometric group (median (IQR), 2.5 (1-4.5)) and isotonic group (median (IQR), 3.0 (2-6))., Conclusions: This is the first study to show a decrease in patellar tendon pain without a modification of training and competition load and the first study to investigate isometric exercises in a clinical setting. Both isometric and isotonic exercise programs are easy-to-use exercises that can reduce pain from patellar tendinopathy for athletes in-season., (Copyright © 2015 Sports Medicine Australia. Published by Elsevier Ltd. All rights reserved.)

The most common methods for attaching a device to the internal tissues of the human body are via sutures, clips or staples. These attachment techniques require penetration and manipulation of the tissue. Tears and leaks can often be a complication post-attachment, and scarring usually occurs around the attachment sites. To resolve these issues, it is proposed to develop a soft tissue scaffold impregnated with Rose Bengal/Chitosan solution (RBC-scaffold, 0.01% w/v Rose Bengal, 1.7% w/v Medium Molecular Weight Chitosan). This scaffold will initially attach to the tissue via a light activation method. The light activates the dye in the scaffold which causes cross-links to form between the scaffold and tissue, thus adhering them together. This is done without mechanically manipulating the surrounding tissue, thus avoiding the issues associated with current techniques. Eventually, the scaffold will be resorbed and tissue will integrate for long-term attachment. A variety of tests were performed to characterise the RBC-scaffold. Porosity, interconnectivity, and mechanical strength were measured. Light activation was performed with a broad spectrum (380-780nm) 10W LED lamp exposed to various time lengths (2-15min, Fluence range 0.4-3J/cm(2) ). Adhesive strength of the light-activated bond was measured with lap-shear tests performed on porcine stomach tissue. Cell culture viability was also assessed to confirm tissue integration potential. These properties were compared to Variotis™, an aliphatic polyester soft tissue scaffold which has proven to be viable for soft tissue regeneration. The RBC-scaffolds were found to have high porosity (86.46±2.95%) and connectivity, showing rapid fluid movement. The elastic modulus of the RBC-scaffolds (3.55±1.28MPa) was found to be significantly higher than the controls (0.15±0.058MPa, p<0.01) and approached reported values for human gastrointestinal tissue (2.3MPa). The maximum adhesion strength achieved of the RBC-scaffolds was 8.61±2.81kPa after 15min of light activation, this is comparable to the adhesion strength of fibrin glue on scaffolds. Cell attachment was seen to be similar to the controls, but cells appeared to have better cell survivability. In conclusion, the RBC-scaffolds show promise for use as a novel light activated attachment device with potential applications in attaching an anti-reflux valve in the lower oesophagus and also in wound healing applications for stomach ulcers., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Previously, we demonstrated that the Rho/ROCK pathway is involved in ouabain-induced apoptosis in HUVEC. In the current work, we investigated whether the Rho/ROCK pathway is functional during cardiac glycosides-induced cytotoxic effects in cancer cell lines, as well as in non-tumor cells. For that purpose, we evaluated the role of ROCK activation in bleb formation and cell migration over upstream and downstream effectors in addition to ROCK cleavage after cardiac glycosides treatment. All three cardiac glycosides (ouabain, digoxin and bufalin) induced cell death in HeLa and HepG2 cells and increased the formation of blebbing in HeLa cells. In contrast to our previous study, ROCK inhibitor Y27632 did not prevent bleb formation. Observation of ROCK II cleavage after ouabain, digoxin and oxaliplatin treatments in HeLa and/or HepG2 cells suggested that cleavage is independent of cell type and cell death induction. While inhibiting cleavage of ROCK II by the caspase inhibitors z-VAD-fmk, z-VDVAD-fmk and z-DEVD-fmk, evaluation of caspase 2 siRNA ineffectiveness on this truncation indicated that caspase-dependent ROCK II cleavage is differentially regulated in cancer cell lines. In HeLa cells, ouabain induced the activation of ROCK, although it did not induce phosphorylation of ERM, an upstream effector. While Y27632 inhibited the migration of HeLa cells, 10nM ouabain had no effect on cell migration. In conclusion, these findings indicate that the Rho/ROCK pathway is regulated differently in cancer cell lines compared to normal cells during cardiac glycosides-induced cell death., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Statins are known to display benefits in various diseases independently from their cholesterol lowering properties. In this study, we investigated the acute effects of atorvastatin on vascular reactivity to various spasmogens in isolated rat aorta. The responses to noradrenaline (NA, 10(-8) -10(-4) m), endothelin-1 (ET-1, 10(-10) -10(-7) m), and potassium chloride (KCl, 10-100 mm) were evaluated in aortic rings pretreated with atorvastatin (10(-7) -10(-4) m, 30 min). To verify the mechanism of action, the effects of atorvastatin were studied in the presence of cholesterol precursor, mevalonate (10(-2) m, 45 min), mevalonate-derived isoprenoids, namely geranylgeranyl pyrophosphate (GGPP, 5 × 10(-6) m, 30 min) and farnesyl pyrophosphate (FPP, 5 × 10(-6) m, 30 min), and in the absence of endothelium. In parallel, aortic rings were pretreated with the specific inhibitor of Rho kinase, Y-27632 (10(-7) -10(-6) m). Atorvastatin significantly and concentration-dependently reduced the contractions to spasmogens in rat aorta. This acute inhibitory effect was also evident in endothelium-denuded rings. Pretreatment with mevalonate and GGPP, but not with FPP, reversed the inhibitory effect of atorvastatin (10(-4) m) on NA and ET-1 induced contractions. Similar to atorvastatin, pretreatment with Y-27632 inhibited the contractions to NA and KCl in a concentration-dependent manner. Western blot analysis revealed that both atorvastatin (10(-4) m) and Y-27632 (10(-6) m) pretreatment inhibited the phosphorylation of myosin phosphatase target subunit-1 (MYPT-1) triggered by NA, indicating an inhibitory influence on myosin phosphatase. In conclusion, atorvastatin displayed an acute inhibitory effect on vascular contractility evoked by various spasmogens and the inhibitory effect was possibly mediated by the inhibition of mevalonate and GGPP synthesis as well as the prevention of MYPT-1 phosphorylation induced by Rho/Rho kinase., (© 2015 Société Française de Pharmacologie et de Thérapeutique.)

Ouabain is an endogenous Na(+)/K(+)-ATPase inhibitor whose chronic administration induces hypertension. Endogenous ouabain levels increase in human essential hypertension. On the other hand, Rho/Rho kinase (ROCK) pathway has been implicated in various animal models of hypertension. In the current work, we evaluated the possible involvement of Rho kinase in ouabain-induced hypertension. Ouabain was administered daily (20 µg/kg, i.p.) to Wistar rats for 6 weeks. After the ouabain treatment, we evaluated the possible changes in vascular responses to KCl and phenylephrine alone and in the presence of Rho kinase inhibitor Y27632. We also determined the expressions of ROCKs, Rho A and phosphorylation of myosin binding subunit of myosin light chain phosphatase (pMYPT) and activation of Rho A. Agonist-induced contractions in the presence of Y27632 are significantly decreased and Y27632-induced relaxations in aortas precontracted with phenylephrine are significantly enhanced with the chronic treatment of ouabain. Although the expressions of ROCK I and ROCK II remained unchanged, pMYPT expression was significantly increased in ouabain-treated group. Moreover, Rho A expression and activation were decreased after treatment with ouabain. Although Rho kinase expression did not change in aortas, increased basal Rho kinase activation may contribute to the development of ouabain-induced hypertension. Our current data present the first evidence that Rho kinase is involved in the development of ouabain-induced hypertension in rats.

Small G protein Rho and its most studied effectors, ROCK I and ROCK II, are involved in several cellular fuctions including smooth muscle and non-muscle cell contractions, cell migration and apoptosis. Activation of ROCK I by caspase-3 and activation of ROCK II by granzyme B are essential for membrane blebbing in the execution phase of apoptosis. In contrast, it has been demonstrated that Rho signaling is critical for blebbing developed after serum removal. As it was shown by us previously, ouabain induces membrane blebbing and proteolitic cleavage of ROCK I and ROCK II via caspases in human umbilical endothelial cells. However, caspase inhibitors do not prevent ouabain-induced blebs. Ouabain induces concentration-dependent cell death and membrane blebbing in endothelial cells. The aim of this study was to identify the possible role of Rho in ouabain-induced membrane blebbing. Pretreatment of endothelial cells with a Rho inhibitor CT04 did not inhibit the ouabain-induced cell death but prevented the development ofbleb formation. These results indicate that bleb formation is dependent on Rho activity in ouabain-induced cell death in HUVECs. Taken together, these results suggest that the mechanism of membrane bleb formation might be different depending on cell type and cell death-stimuli.

The aim of our study was to investigate the effect of taurine on the relationship between nitric oxide (NO), asymmetric dimethylarginine (ADMA) and homocysteine (Hcy) in endotoxin-induced human umblical vein endothelial cell (HUVEC) cultures. For this reason, four groups were formed (n=12). Control group consists of HUVEC cultures without any treatment. Lipopolysaccharide (LPS) and LPS+taurine groups were treated with 10 μg/mL endotoxin, 5 μg/mL taurine and endotoxin+taurine (same doses), respectively. Nitrite/nitrate (NOx), ADMA and Hcy levels were measured. There was a significant increase of NOx, ADMA and Hcy in endotoxemia (p<0.05). Taurine treatment elevated NOx levels significantly (p<0.01) in taurine and LPS + taurine group compared to control group, while it reduced NOx levels compared to LPS group. In contrast, taurine decreased ADMA levels to the control level both in taurine and taurine+LPS group compared to LPS. Hcy levels increased significantly compared to taurine group (p<0.05) and did not change compared to LPS group. Taurine was effective on ADMA-NO relationship whereas no beneficial effect was observed in Hcy levels (p<0.05).

Background: Lateral Epicondylalgia (LE) is a common injury for which no reliable and valid measure exists to determine severity in the Dutch language. The Patient-Rated Tennis Elbow Evaluation (PRTEE) is the first questionnaire specifically designed for LE but in English. The aim of this study was to translate into Dutch and cross-culturally adapt the PRTEE and determine reliability and validity of the PRTEE-D (Dutch version)., Methods: The PRTEE was cross-culturally adapted according to international guidelines. Participants (n = 122) were asked to fill out the PRTEE-D twice with a one week interval to assess test-retest reliability. Internal consistency of the PRTEE-D was determined by calculating Crohnbach's alphas for the questionnaire and subscales. Intraclass Correlation Coefficients (ICC) were calculated for the overall PRTEE-D score, pain and function subscale and individual questions to determine test-retest reliability. Additionally, the Disabilities for the Arm, Shoulder and Hand questionnaire (DASH) and Visual Analogue Scale (VAS) pain scores were obtained from 30 patients to assess construct validity; Spearman's correlation coefficients were calculated between the PRTEE-D (subscales) and DASH and VAS-pain scores., Results: The PRTEE was successfully cross-culturally adapted into Dutch (PRTEE-D). Crohnbach's alpha for the first assessment of the PRTEE-D was 0.98; Crohnbach's alpha was 0.93 for the pain subscale and 0.97 for the function subscale. ICC for the PRTEE-D was 0.98; subscales also showed excellent ICC values (pain scale 0.97 and function scale 0.97). A significant moderate correlation exists between PRTEE-D and DASH (0.65) and PRTEE-D and VAS pain (0.68)., Conclusion: The PRTEE was successfully cross-culturally adapted and this study showed that the PRTEE-D is reliable and valid to obtain an indication of severity of LE. An easy-to-use instrument for practitioners is now available and this facilitates comparing Dutch and international research data.

In this study, we synthesized some novel N-(tetrazol-1H-5-yl)-6,14-endoethenotetrahydrothebaine 7α-substituted 1,3,4-oxadiazole and 1,3,4-thiadiazole derivatives as potential analgesic agents. The structures of the compounds were established on the basis of their IR, ¹H NMR, ¹³C NMR, 2D NMR, and high-resolution mass spectral data. The analgesic activity was evaluated by a rat-hot plate test model and a rat tail-flick model. Compound 12 showed analgesic activity higher than that of morphine. In addition to a histopathological and biochemical evaluation, the LD₅₀ dose for the most active compound 12 was determined., (Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Indirect acute lung injury is associated with high morbidity and mortality. We investigated the link between Rho kinase (ROCK) activation and apoptotic cell death in sepsis induced acute lung injury. This hypothesis was tested by administering a specific, selective inhibitor of ROCK (Y-27632) to rats subjected to cecal ligation and puncture (CLP). Rats were randomly divided into 4 groups as; sham-operated, sham + Y-27632, CLP and CLP + Y-27632. Twenty-four hours later, each experiment was terminated and lungs analyzed. Histopathology was assessed by hematoxylin-eosin staining and the presence of apoptosis was evaluated through the TUNEL assay. Pulmonary activity of caspase 3 and ROCK 1 & 2 were measured by western blot. Interstitial edema, severely damaged pulmonary architecture with massive infiltration of the inflammatory cells and an increase in lung tissue TBARS levels as well as 3-NT to total tyrosine ratios were observed in untreated CLP animals. Pretreatment of animals with Y-27632, reduced lung injury in the CLP induced septic rats in each of these parameters of lung injury (p<0.05). Western immunoblot revealed active caspase cleavage and increased expression of active fragment of ROCK 1 & 2 in the CLP group. TUNEL assay showed an increase in percentage of apoptotic cells when comparing the CLP group with the CLP + Y-27632 group. These results suggest an important role of Rho kinase in sepsis induced lung injury by a mechanism that might be related to oxidative and/or nitrosative stress mediated caspase cleavage leading to apoptosis.

Ouabain, a specific Na+/K+-ATPase inhibitor, has recently been identified as a mammalian hormone. Its elevated concentrations in human plasma have also been associated with pathogenesis of several diseases. Recent studies have shown that ouabain induces aponecrotic cell death in a cell-type- and dose-dependent manner. However, the exact mechanism of ouabain-induced cell death is not fully understood. The Rho GTPase effectors Rho kinases-1 and -2 (Rock-1 and Rock-2) which play central roles in the organization of the actin cytoskeleton, involve in several models of apoptosis. In this study, we investigated the possible involvement of Rocks in ouabain-induced human umbilical vein endothelial cell (HUVEC) apoptosis. Ouabain treatment resulted in loss of cell-cell and cell-substratum adhesion and apoptotic blebbing. Pretreatment of cells with Y-27632, a specific Rock inhibitor, resulted in the inhibition of cell-to-cell detachment and formation of membrane blebs. However, Y-27632 did not prevent ouabain-induced cell-substratum detachment. Instead, treatment with Y-27632 actually accelerated this process. Ouabain treatment induced cleavage of Rock-1 and Rock-2, which was prevented by caspase-3 and caspase-2 specific inhibitors z-DEVD-fmk and z-VDVAD-fmk, respectively. Ouabain-induced Rock-2 cleavage generated a fragment of approximately 140 kDa corresponding to the consensus sequence of caspase-2 on the carboxy terminus of Rock-2. Although it has been previously shown that Rock-2 was cleaved by caspase-2, we have identified here a novel cleavage site and fragment of Rock-2. Our data indicate that ouabain induces both Rock-1 and Rock-2 cleavage via caspase-dependent mechanisms and provide evidence that Rocks are involved in ouabain-induced cell-to-cell detachment and apoptosis.

Short ischemic episodes increase tolerance against subsequent severe ischemia in the heart. Nitropropionate (3-NP), an irreversible inhibitor of succinic dehydrogenase of the mitochondrial complex II, was shown to induce protective effect against ischemic brain injury. The aim of this study was to investigate the possible protective effect of 3-NP on regional ischemia in preconditioned rat heart in vivo. Hearts were assigned into three groups: first, in order to induce ischemic preconditioning (IP) 5 min ischemia separated by 10 min reperfusion protocol was used; second, non-preconditioned group was used as control; and third, 3-NP (20 mg/kg, i.p.) was injected 3 h before the surgical procedure in order to induce chemical preconditioning. In all these groups, 30 min regional ischemia was followed by 60 min reperfusion. Infarct size, bax expression, number of ventricular ectopic beats (VEB), duration of ventricular tachycardia (VT) and ventricular fibrillation (VF) were significantly decreased in ischemic preconditioning and 3-NP pretreatment groups, whereas bcl-2 values were not markedly changed in these groups during occlusion period. These results showed that in the anesthetized rat heart 3-NP induced chemical preconditioning by decreasing infarct size, number of VEB, duration of VT and VF. Protective effect is associated with via decreased production of bax protein expression.

Rho kinase (ROCK) and nitric oxide (NO) are important targets in cardiovascular diseases. Therefore, we investigated the possible influence of NO on Rho kinase (ROCK-2 isoform) expressions in cultured rat coronary microvascular endothelial cells. The cells were isolated from Wistar rats on a Langendorff system, and were incubated overnight (approximately 16 h) with an NO generator, A-23187 (10 to 10 M), NO donors, such as sodium nitroprusside (10 to 10 M), glyceryl trinitrate (10 to 10 M), 2,2'-(hydroxynitrosohydrazono)bis-ethanimine (10 to 10 M), and NaNO2 (10 to 10 M) or a nitric oxide synthase (NOS) inhibitor, N-nitro-L-arginine methylester (2 x 10 M), or two ROCK inhibitors, (+)-(R)-trans-4-(1-aminoethyl)- N-(4-pyridyl) cyclohexanecarboxamide dihydrochloride monohydrate (Y-27632, 10 M) and fasudil (10 M) in the absence or presence of thrombin (4 U/mL). ROCK-2 and endothelial NOS (eNOS) expressions were detected by Western blotting. Moreover, nitrite/nitrate levels were detected by Griess method in the presence of the ROCK inhibitors. The NO donors and the NO generator had no significant effects on ROCK-2 expression. Y-27632 and fasudil did not alter eNOS expression and NO production. Nitrite/nitrate levels were 4.4 +/- 0.32 microM in control and 4.0 +/- 0.93 microM and in Y-27632 group. These results demonstrate that prolong NO donation could not suppress the expression of ROCK-2 protein, and the ROCK inhibitor did not change e-NOS expression and NO production in the cultured rat coronary microvascular endothelial cells.