CAMSAP2 enhances lung cancer cell metastasis by mediating RASAL2 degradation.

AIMS: Cancer metastasis significantly contributes to mortality in lung cancer patients. Calmodulin-regulated spectrin-associated protein family member 2 (CAMSAP2) plays a significant role in cancer cell migration; however, its role in lung cancer metastasis and the underlying mechanism remain largely unknown. The present study aimed to investigate the impact of CAMSAP2 on lung cancer. MAIN METHODS: The clinical relevance of CAMSAP2 in lung cancer patients was assessed using public database. RNA interference experiments were conducted to investigate role of CAMSAP2 in cell migration through transwell and wound healing assays. Molecular mechanisms were explored by identifying the possible interacting partners and pathways using the BioGRID and KEGG pathway analyses. The impact of CAMSAP2 on Ras protein activator-like 2 (RASAL2)-mediated lung cancer metastasis was investigated through biochemical assays. Additionally, in vivo experimentation using a murine tail vein metastasis model was performed to comprehend CAMSAP2's influence on metastasis. KEY FINDINGS: A high expression level of CAMSAP2 was associated with poor overall survival in lung cancer patients and it positively correlated with cell migration in non-small cell lung cancer (NSCLC) cell lines. Knockdown of CAMSAP2 inhibited lung cancer cell motility in vitro and metastasis in vivo. Proteomic and biochemical analyses revealed the interaction between CAMSAP2 and RASAL2, which facilitates the degradation of RASAL2 through the ubiquitin-proteasome system. These degradation processes resulted in the activation of the extracellular signal-regulated kinase (ERK) signaling pathway, thereby promoting lung cancer metastasis. Collectively, the results of this study suggest that CAMSAP2 is a crucial regulator of cancer cell migration and metastasis and a promising therapeutic target for lung cancer.

Preclinical Characterization of 22-(4'-Pyridinecarbonyl) Jorunnamycin A against Lung Cancer Cell Invasion and Angiogenesis via AKT/mTOR Signaling.

Non-small-cell lung cancer (NSCLC), the most prevalent form of lung cancer, is associated with an unfavorable prognosis owing to its high rate of metastasis. Thus, the identification of new drugs with potent anticancer activities is essential to improve the clinical outcome of this disease. Marine organisms exhibit a diverse source of biologically active compounds with anticancer effects. The anticancer effects of jorunnamycin A (JA) derived from the Thai blue sponge (Xestospongia sp.) and 22-(4'-pyridinecarbonyl) jorunnamycin A (22-(4'-py)-JA), the semisynthetic derivative of JA, have been reported. The present study aimed to investigate the impact of 22-(4'-py)-JA on NSCLC metastasis using in vitro, in vivo, and in silico approaches. The JA derivative inhibited tumor cell invasion and tube formation in human umbilical vein endothelial cells (HUVECs). The computational analysis demonstrated strong and stable interactions between 22-(4'-py)-JA and the AKT protein. Further examinations into the molecular mechanisms revealed the suppression of AKT/mTOR/p70S6K signaling by 22-(4'-py)-JA, leading to the downregulation of matrix metalloproteinases (MMP-2 and MMP-9), hypoxia-inducible factor-1α (HIF-1α), and vascular endothelial growth factor (VEGF). Furthermore, 22-(4'-py)-JA suppressed in vivo metastasis by decreasing the number of colonies in the lung. These findings indicated the antimetastasis activity of 22-(4'-py)-JA, which might prove useful for further clinical applications.

Interpenetrating Low-Molecular Weight Hyaluronic Acid in Hyaluronic Acid-Based In Situ Hydrogel Scaffold for Periodontal and Oral Wound Applications.

Tissues engineering has gained a lot of interest, since this approach has potential to restore lost tooth-supporting structures, which is one of the biggest challenges for periodontal treatment. In this study, we aimed to develop an in situ hydrogel that could conceivably support and promote the regeneration of lost periodontal tissues. The hydrogel was fabricated from methacrylated hyaluronic acid (MeHA). Fragment/short-chain hyaluronic acid (sHA) was incorporated in this hydrogel to encourage the bio-synergistic effects of two different molecular weights of hyaluronic acid. The physical properties of the hydrogel system, including gelation time, mechanical profile, swelling and degrading behavior, etc., were tested to assess the effect of incorporated sHA. Additionally, the biological properties of the hydrogels were performed in both in vitro and in vivo models. The results revealed that sHA slightly interfered with some behaviors of networking systems; however, the overall properties were not significantly changed compared to the base MeHA hydrogel. In addition, all hydrogel formulations were found to be compatible with oral tissues in both in vitro and in vivo models. Therefore, this HA-based hydrogel could be a promising delivery system for low molecular weight macromolecules. Further, this approach could be translated into the clinical applications for dental tissue regeneration.

In Vivo Biocompatible Self-Assembled Nanogel Based on Hyaluronic Acid for Aqueous Solubility and Stability Enhancement of Asiatic Acid.

Asiatic acid (AA), a natural triterpene found in Centalla asiatica, possesses polypharmacological properties that can contribute to the treatment and prophylaxis of various diseases. However, its hydrophobic nature and rapid metabolic rate lead to poor bioavailability. The aim of this research was to develop a thermoresponsive nanogel from hyaluronic acid (HA) for solubility and stability enhancement of AA. Poly(N-isopropylacrylamide) (pNIPAM) was conjugated onto HA using a carbodiimide reaction followed by 1H NMR characterization. pNIPAM-grafted HA (HA-pNIPAM) nanogels were prepared with three concentrations of polymer, 0.1, 0.15 and 0.25% w/v, in water by the sonication method. AA was loaded into the nanogel by the incubation method. Size, morphology, AA loading capacity and encapsulation efficiency (EE) were analyzed. In vitro cytocompatibility was evaluated in fibroblast L-929 cells using the PrestoBlue assay. Single-dose toxicity was studied using rats. HA-pNIPAM nanogels at a 4.88% grafting degree showed reversible thermo-responsive behavior. All nanogel formulations could significantly increase AA water solubility and the stability was higher in nanogels prepared with high polymer concentrations over 180 days. The cell culture study showed that 12.5 µM AA in nanogel formulations was considered non-toxic to the L-929 cells; however, a dose-dependent cytotoxic effect was observed at higher AA-loaded concentrations. In vivo study proved the non-toxic effect of AA loaded in HA-pNIPAM nanogels compared with the control. Taken together, HA-pNIPAM nanogel is a promising biocompatible delivery system both in vitro and in vivo for hydrophobic AA molecules.

Tiletamine/zolazepam and dexmedetomidine with tramadol provide effective general anesthesia in rats.

Background: Tiletamine/zolazepam is a dissociative anesthetic combination commonly used in small animals but information is limited in rats. The alpha-2 agonist, dexmedetomidine, has gained popularity in laboratory animal anesthesia. Tramadol is a weak opioid mu agonist. The aim of this study was to assess whether the tiletamine/zolazepam/dexmedetomidine (ZD) combination effectively provides a surgical anesthesia plane comparable to tiletamine/zolazepam/dexmedetomidine with tramadol (ZDT) in a minor procedure in rats. Methods: Rats were induced with ZD or ZDT. After the loss of paw withdrawal, a small incision was made on the rats' left thighs as a surgical stimulus. Rats were maintained under a surgical anesthesia plane by assessing the loss of the paw withdrawal reflex for 45 minutes, then atipamezole was administered. Monitored anesthesia parameters included: (a) physiological parameters - pulse rate (PR), respiratory rate (RR), tissue oxygen saturation (%SpO2), and body temperature; (b) duration parameters - induction time, onset and duration of surgical anesthesia plane, onset of recovery, and recovery time. Results: PR was significantly lower at 10 minutes in ZD and 5 minutes in ZDT groups. No difference was observed for RR, %SpO2, and body temperature. Likewise, there were no differences for duration parameters: induction time was less than 3 minutes; onset and duration of surgical anesthesia plane were approximately 5 and 45 minutes, respectively; onset of recovery (time to move) was 51 minutes; and recovery time was 52 minutes, respectively. Conclusion: These data suggest the ZD combination provides a surgical anesthesia plane comparable to ZDT in a rat incisional pain model.

Acute effect of ivabradine on heart rate and myocardial oxygen consumption in dogs with asymptomatic mitral valve degeneration.

Degenerative mitral valve disease (DMVD) is a common cardiac disease in geriatric dogs characterized by the degeneration of the mitral valve, leading to decreased cardiac output and activation of the sympathetic and renin-angiotensin-aldosterone system. This disease results in an increased resting heart rate (HR) and myocardial oxygen consumption (MVO2). A recent publication demonstrated that dogs with asymptomatic DMVD had a significantly higher HR and systemic blood pressure (BP) than age-matched control dogs. This higher HR will eventually contribute to increased MVO2. This study aimed to determine the effects of a single oral dose of ivabradine on the HR, MVO2 as assessed by the rate-pressure product, and BP in dogs with asymptomatic DMVD. Seven beagles with naturally occurring DMVD were instrumented by the Holter recorder and an oscillometric device to measure electrocardiogram and BP for 24 and 12 h, respectively. Each dog was randomly subjected to receive either placebo or ivabradine (0.5, 1.0 and 2.0 mg/kg). The results revealed that oral administration of ivabradine significantly decreased the HR and rate-pressure product in a dose-dependent manner without adverse effects. The highest dose of 2.0 mg/kg significantly reduced systolic and mean BP. Therefore, the findings imply that a single oral ivabradine administration at a dose of 1.0 mg/kg is suitable for dogs with asymptomatic DMVD to reduce the HR and MVO2 without marked effects on BP. This may potentially make ivabradine promising for management of an elevated HR in DMVD dogs.

An increasing electromechanical window is a predictive marker of ventricular fibrillation in anesthetized rabbit with ischemic heart.

The QTc interval is widely used in Safety Pharmacological studies to predict arrhythmia risk, and the electromechanical window (EMW) and short-term variability of QT intervals (STVQT) have been studied as new biomarkers for drug-induced Torsades de Pointes (TdP). However, the use of EMW and STVQT to predict ventricular fibrillation (VF) has not been elucidated. This study aimed to evaluate EMW and STVQT to predict VF in anesthetized rabbit model of VF. VF was induced by ligation of the left anterior descending and a descending branch of the left circumflex coronary arteries in a sample population of rabbits (n=18). VF was developed 55.6% (10/18). In rabbit with VF, the EMW was significantly higher than in rabbits without VF (96.3 ± 15.6 ms and 49.5 ± 5.6 ms, respectively, P<0.05). STVQT had significantly increased before the onset of VF in rabbits that experienced VF, but not in rabbits that did not experience VF (11.7 ± 1.8 ms and 3.7 ± 0.4 ms, respectively, P<0.05). The EMW and STVQT had better predictive power for VF with higher sensitivity and specificity than the QTc measure. The result suggested that the increasing of EMW, as well as the elevation of STVQT, can potentially be used as biomarkers for predicting of VF.

Characteristics of electromechanical window in anesthetized rabbit models of short QT and long QT syndromes.

The current regulatory guidelines recommend the use of QT interval to assess the risk of arrhythmogenic potential of new chemical entities. Recently, the electromechanical window (EMW), the difference in duration between electrical and mechanical systole, has been proposed as markers for drug-induced torsades de pointes (TdP); however, data of EMW in short QT model are not available. This study aimed to characterize the EMW as a marker for drug-induced ventricular arrhythmias in anesthetized rabbit model of long QT syndrome type 2 (LQT2) and short QT syndrome (SQTS) infused with reference compounds known to lengthen or shorten QT intervals. After rabbits were anesthetized with isoflurane, body surface electrocardiograms and left ventricular pressure were recorded. The LQT2 was produced by intravenous infusion with dofetilide (n = 6), quinidine (n = 6) and sotalol (n = 6) whereas the SQTS was induced by intravenous escalating concentrations of nicorandil (n = 7), pinacidil (n = 5) and cromakalim (n = 5). The EMW in anesthetized rabbits ranged from 1.3 to 53.3 msec. All three drugs known to lengthen QT intervals prolonged QT and QTcF interval while the EMW was markedly decreased to negative values. Pinacidil significantly produced QT and QTcF shortening and significantly abbreviated the EMW (p < 0.05). This study demonstrated that the EMW is associated with QT intervals (p < 0.001). It is negative in the presence of QT-prolonging drugs while it is more positive in the presence of QT-shortening drugs. The results suggest that the EMW in anesthetized rabbits can be used in drug safety evaluation in addition to the QT interval.

Sildenafil improves heart rate variability in dogs with asymptomatic myxomatous mitral valve degeneration.

Myxomatous mitral valve degeneration (MMVD) causes an imbalance of sympathovagal activity resulted in poor cardiac outcomes. Phosphodiesterase-5 inhibitors have been revealed cardioprotective effect in patients with heart diseases. This study aimed to 1) compare the heart rate variability (HRV) between asymptomatic MMVD and healthy dogs and 2) assess long-term effects of sildenafil and enalapril on time- and frequency-domains analyzes. Thirty-four dogs with MMVD stage B1 or B2 and thirteen healthy dogs were recruited into the study. MMVD dogs were divided into 3 subgroups: control (n=13), sildenafil (n=12) and enalapril (n=9). HRV was analyzed from 1-hr Holter recording at baseline (D0) in all dogs and at 30, 90 and 180 days after treatment. The results showed that MMVD dogs had significant higher heart rate (HR), systemic blood pressures, the ratio of low to high frequency (LF/HF) and had significant decreased standard deviation of all normal to normal RR intervals (SDNN) and the percentage of the number of normal-to-normal sinus RR intervals with differences >50 msec computed over the entire recording (pNN50) when compared with healthy dogs (P<0.05). Neither time nor frequency domain parameters were different among subgroups of MMVD dogs at D0. After treatment with sildenafil for 90 days, both time- and frequency-domain parameters were significantly increased when compared with control and enalapril groups. This study demonstrated that sildenafil improves HRV in asymptomatic MMVD dogs suggesting that sildenafil should be used in the MMVD dogs to restore the sympathovagal balance.

Dronedarone attenuates the duration of atrial fibrillation in a dog model of sustained atrial fibrillation.

Atrial fibrillation (AF) is a supraventricular arrhythmia that leads to a decrease in cardiac output and impairs cardiac function and quality of life. Dronedarone has an atrial-selective property and has been used for management of AF in humans, but limited information is available in dogs. This study was designed to evaluate efficacy of dronedarone in attenuating the duration of AF in dog model of sustained AF. Six beagle dogs were anesthetized with isoflurane and instrumented to measure atrial action potential duration (aAPD) and atrial effective refractory period (AERP). Then AF was induced by rapid right atrial pacing (20 V, 40 Hz) simultaneously with infusion of phenylephrine (2 µg/kg/min, intravenously) for 20 min. The duration of sustained AF was recorded, and the animals were allowed to recover. Dronedarone was given at a dose of 20 mg/kg, BID, orally for 7 days. On the last day, the dogs were anesthetized again to record aAPD and AERP, and AF was induced with the same procedure as described above. The results showed that after dronedarone administration the aAPD was lengthened significantly from 76.4 ± 4.2 ms to 91.2 ± 3.9 ms (P<0.05) and AERP was prolonged significantly from 97.5 ± 2.8 ms to 120 ± 4.8 ms (P<0.05). The duration of sustained AF was also significantly attenuated after receipt of dronedarone (P<0.05). It can be suggested that oral dronedarone attenuates the duration of sustained AF in a dog model of AF by extending the AERP more than the aAPD, causing post-repolarization refractoriness. Hence, dronedarone may be useful for management of AF in dogs.

Acute effects of intravenous dronedarone on electrocardiograms, hemodynamics and cardiac functions in anesthetized dogs.

Dronedarone is a class III antiarrhythmic that has been used for management of atrial fibrillation in humans, but limited information was found in dogs. The objective of this study was to determine the acute effects of escalating concentrations of dronedarone on electrocardiograms (ECG), hemodynamics and cardiac mechanics in healthy dogs. A total of 7 beagle dogs were anesthetized with isoflurane and instrumented to obtain lead II ECG, pressures at ascending aorta, right atrium, pulmonary artery and left ventricle, and left ventricular pressure-volume relationship. Five dogs were given vehicle and followed by escalating doses of dronedarone (0.5, 1.0 and 2.5 mg/kg, 15 min for each dose), and two dogs were used as a vehicle-treated control. All parameters were measured at 15 min after the end of each dose. The results showed that all parameters in vehicle-treated dogs were unaltered. Dronedarone at 2.5 mg/kg significantly lengthened PQ interval (P<0.01), reduced cardiac output (P<0.01) and increased systemic vascular resistance (P<0.01). Dronedarone produced negative inotropy assessed by significantly lowered end-systolic pressure-volume relationship, preload recruitable stroke work, contractility index and dP/dtmax. It also impaired diastolic function by significantly increased end-diastolic pressure-volume relationship, tau and dP/dtmin. These results suggested that acute effects of dronedarone produced negative dromotropy, inotropy and lusitropy in anesthetized dogs. Care should be taken when given dronedarone to dogs, especially when the patients have impaired cardiac function.

Transmural dispersion of repolarization and cardiac remodeling in ventricles of rabbit with right ventricular hypertrophy.

INTRODUCTION: Recent publications demonstrated that rabbits with right ventricular hypertrophy (RVH) possess high sensitivity and specificity for drug-induced arrhythmias. However, the underlying mechanism has not been elucidated. This study aimed to evaluate RVH induced changes in cardiac remodeling especially the transmural dispersion of repolarization (TDR), epicardial monophasic action potentials (MAP), and hERG mRNA expression in rabbits. METHODS: New Zealand White rabbits (n=13) were divided into 2 groups: sham operated (SHAM, n=6) and pulmonary artery banding (PAB, n=7). PAB was induced by narrowing the pulmonary artery. Twenty weeks after surgery, hemodynamic, cardiac function, electrocardiograms, and MAP were obtained from PAB compared with SHAM. After measurement, rabbits were sacrificed to collect ventricular myocardium for histopathological analysis and measurement of hERG mRNA expression by real time PCR. RESULTS: After 20weeks, the % HW to BW ratio of whole heart and right ventricle (RV) and left and right ventricular free wall thickness was significantly increased in PAB when compared with those in SHAM. PAB has a significant electrical remodeling as demonstrated by lengthening of QT, QTc intervals, and increased Tp-Te duration. PAB also has a significant functional remodeling verified by decreased contractility index of RV and lengthened time constant of relaxation of LV. MAP of RV epicardium was significantly shortened in PAB consistently with increased hERG mRNA expression at the epicardium of RV. DISCUSSION: The rabbit with PAB demonstrates cardiac remodeling diastolic and systolic dysfunctions. These rabbits also demonstrate increased TDR and electrical remodeling related to the change of hERG mRNA expression which may be prone to develop arrhythmias.