Ultrasound-guided selective supraclavicular nerve block for postoperative pain control in children receiving Hickman catheter or chemoport insertion: A randomized controlled trial.

BACKGROUND: Optimal pain management after insertion of a central venous catheter in children remains unclear. AIM: This study aimed to evaluate the effects of a selective supraclavicular nerve block on postoperative analgesia in pediatric patients undergoing hickman catheter or chemoport insertion. METHODS: Fifty patients aged 3-18 years scheduled for elective Hickman or chemoport insertion were randomized into two groups of 25 each: one group received an ultrasound-guided selective supraclavicular nerve block with 0.1 mL/kg of 0.5% ropivacaine (SSCNB group), and the other group did not receive a nerve block (control group). The primary outcome was the postoperative Wong-Baker Faces Pain Rating Scale score measured between 10 and 30 min after surgery. Secondary outcomes included pain scores at 1, 3, and 24 h after the surgery, block-related complications, length of stay in the postanesthesia care unit, postoperative analgesic consumption, and time to first analgesic use 24 h after surgery. RESULTS: The worst pain score within 30 min in the recovery room was significantly lower in the SSCNB group compared to the control group (6 [5-7] vs. 3 [2-4]; median difference, -3; 95% CI, -4 to -1; p < .001). Pain scores at 1, 3, and 24 h after surgery were also significantly lower in the SSCNB group. The need for both opioid and non-opioid analgesics in the postoperative period was significantly lower in the SSCNB group (36.0% vs. 0%; p = .002 and 44.0% vs. 16.0%; mean difference, -28%; 95% CI, -56 to 0.19; p = .033, respectively), while other secondary outcomes were not significantly different between the two groups. CONCLUSIONS: Ultrasound-guided SSCNB is an effective method for managing postoperative pain in children undergoing Hickman catheter or chemoport insertion, reducing the need for analgesics within 24 h after surgery.

Driving pressure-guided ventilation and postoperative pulmonary complications in thoracic surgery: a multicentre randomised clinical trial.

BACKGROUND: Airway driving pressure, easily measured as plateau pressure minus PEEP, is a surrogate for alveolar stress and strain. However, the effect of its targeted reduction remains unclear. METHODS: In this multicentre trial, patients undergoing lung resection surgery were randomised to either a driving pressure group (n=650) receiving an alveolar recruitment/individualised PEEP to deliver the lowest driving pressure or to a conventional protective ventilation group (n=650) with fixed PEEP of 5 cm H2O. The primary outcome was a composite of pulmonary complications within 7 days postoperatively. RESULTS: The modified intention-to-treat analysis included 1170 patients (mean [standard deviation, sd]; age, 63 [10] yr; 47% female). The mean driving pressure was 7.1 cm H2O in the driving pressure group vs 9.2 cm H2O in the protective ventilation group (mean difference [95% confidence interval, CI]; -2.1 [-2.4 to -1.9] cm H2O; P<0.001). The incidence of pulmonary complications was not different between the two groups: driving pressure group (233/576, 40.5%) vs protective ventilation group (254/594, 42.8%) (risk difference -2.3%; 95% CI, -8.0% to 3.3%; P=0.42). Intraoperatively, lung compliance (mean [sd], 42.7 [12.4] vs 33.5 [11.1] ml cm H2O-1; P<0.001) and Pao2 (median [inter-quartile range], 21.5 [14.5 to 30.4] vs 19.5 [13.5 to 29.1] kPa; P=0.03) were higher and the need for rescue ventilation was less frequent (6.8% vs 10.8%; P=0.02) in the driving pressure group. CONCLUSIONS: In lung resection surgery, a driving pressure-guided ventilation improved pulmonary mechanics intraoperatively, but did not reduce the incidence of postoperative pulmonary complications compared with a conventional protective ventilation. CLINICAL TRIAL REGISTRATION: NCT04260451.

The effect of dexmedetomidine on neuroprotection in pediatric cardiac surgery patients: study protocol for a prospective randomized controlled trial.

BACKGROUND: Infants undergoing cardiac surgery under cardiopulmonary bypass are vulnerable to postoperative neurodevelopmental delays. Dexmedetomidine has been shown to have protective effects on the heart, kidneys, and brain in animals and adults undergoing cardiac surgery with cardiopulmonary bypass. We hypothesized that dexmedetomidine would have a neuroprotective effect on infants undergoing cardiopulmonary bypass and planned a prospective randomized controlled trial with postoperative neurodevelopment measurements. METHODS: This is a single-center, prospective, double-blinded, randomized controlled trial with 1:1 allocation. A cohort of 160 infants undergoing cardiac surgery with cardiopulmonary bypass will be enrolled. After induction, dexmedetomidine will be infused with a loading dose of 1 µg/kg and a maintenance dose of 0.5 µg/kg/h or the same amount of normal saline will be administered. Upon initiation of cardiopulmonary bypass, an additional dose of dexmedetomidine (0.01 µg/cardiopulmonary priming volume) will be mixed with the cardiopulmonary bypass circuit. The primary outcome will be the proportion of infants who score lower than 85 in any of the cognitive, language, or motor Bayley scales of infant development-III tests 1 year after the surgery. Other feasible outcome measures will include differences in plasma glial fibrillary acidic protein, troponin I, interleukin-6, urinary neutrophil gelatinase-associated lipocalin, and perioperative major adverse events. The results of the Bayley scales of infant development-III test from the study group and the control group will be compared using a chi-squared test under intention-to-treat analysis. A generalized estimating equation will be used to analyze repeated measurements over time. DISCUSSION: This study will enable us to assess whether the use of dexmedetomidine can alter the early neurodevelopmental outcome in infants undergoing cardiac surgery with cardiopulmonary bypass and also estimate effects of dexmedetomidine on other organs. TRIAL REGISTRATION: ClinicalTrials.gov NCT04484922. Registered on 24 July 2020.

Gene Network Analysis for Osteoporosis, Sarcopenia, Diabetes, and Obesity in Human Mesenchymal Stromal Cells.

The systemic gene interactions that occur during osteoporosis and their underlying mechanisms remain to be determined. To this end, mesenchymal stromal cells (MSCs) were analyzed from bone marrow samples collected from healthy individuals (n = 5) and patients with osteoporosis (n = 5). A total of 120 osteoporosis-related genes were identified using RNA-sequencing (RNA-seq) and Ingenuity Pathway Analysis (IPA) software. In order to analyze these genes, we constructed a heatmap of one-way hierarchical clustering and grouped the gene expression patterns of the samples. The MSCs from one control participant showed a similar expression pattern to that observed in the MSCs of three patients with osteoporosis, suggesting that the differentiating genes might be important genetic determinants of osteoporosis. Then, we selected the top 38 genes based on fold change and expression, excluding osteoporosis-related genes from the control participant. We identified a network among the top 38 genes related to osteoblast and osteoclast differentiation, bone remodeling, osteoporosis, and sarcopenia using the Molecule Activity Predictor program. Among them, 25 genes were essential systemic genes involved in osteoporosis. Furthermore, we identified 24 genes also associated with diabetes and obesity, among which 10 genes were involved in a network related to bone and energy metabolism. The study findings may have implications for the treatment and prevention of osteoporosis.

Validation of the Masimo O3™ regional oximetry device in pediatric patients undergoing cardiac surgery.

We assessed the accuracy of Masimo O3™ regional cerebral oxygen saturation (rSO2) readings by comparing them with reference values and evaluated the relationship between rSO2 and somatic tissue oxygen saturation (StO2) in children undergoing cardiac surgery. After anesthesia induction, pediatric sensors were applied to the forehead and foot sole, and rSO2 and StO2 values were monitored continuously. Before cardiopulmonary bypass (CPB), FIO2 was set to 0.2, 0.5, and 0.8 serially every 15 min. After CPB, FIO2 was reversed. The reference values (SavO2) were calculated by combining arterial (SaO2) and central venous oxygen saturation (SvO2) readings from the arterial and central lines, respectively (0.7 [Formula: see text] SvO2 + 0.3 [Formula: see text] SaO2). In total, 265 pairs of rSO2/StO2 and SavO2 from 49 patients were analyzed. The bias, standard deviation (SD), standard error (SE), and root mean squared error (RMSE) of rSO2 were 2.6%, 4.5%, 0.3%, and 4.3%, respectively. The limits of agreement ranged from -6.3% to 11.6%. Trend accuracy analysis yielded a relative mean error of -1.4%, with an SD of 4.3%, SE of 0.2%, and RMSE of 3.9%. According to multiple linear regression analysis, the application of CPB, FIO2, Hb level, and tip location of the central venous catheter influenced the bias (all P < 0.05). Furthermore, the correlation between rSO2 and StO2 was weak (r = 0.254). rSO2 readings by the Masimo O3™ device and pediatric sensor had good absolute and trending accuracies with respect to the calculated reference values in children undergoing cardiac surgery. rSO2 and StO2 cannot be used interchangeably.Clinical trial registration http://clinicaltrials.gov (number: NCT04208906).

Central venous catheter-related thrombosis in pediatric surgical patients: A prospective observational study.

BACKGROUND: Perioperative central venous catheters are required but may be associated with various complications. AIMS: The purpose of our study was to assess the incidence and perioperative risk factors for catheter-related internal jugular vein thrombosis in pediatric surgical patients. METHODS: This prospective observational study included children under 6 years of age who were scheduled to undergo central venous catheterization of the right internal jugular vein under general anesthesia. A central venous catheter was inserted under real-time ultrasound guidance. An investigator examined for thrombosis using ultrasonography at predetermined time points. The primary aim was the incidence of catheter-related thrombosis from insertion until the 5th day postoperatively or the removal of the central venous catheter. The secondary aim was the determination of the risk factors for thrombosis. RESULTS: Eighty patients completed the study. Internal jugular vein thrombi were found in 31 patients (38.8%, 95% CI 28.0-49.4). On multiple logistic regression analyses, the number of insertion attempts was the only influencing factor for catheter-related thrombosis (p < .001). More than two insertion attempts increased the risk of thrombosis (odds ratio 5.6; 95% CI 1.7 - 18.7, p = .004). Anesthesia time (p = .017; mean difference 166.4 min; 95% CI 55.7-277.1), intraoperative red blood cell transfusion (p = .001; median difference 21.1 ml kg-1 ; 95% CI 6.6-34.4), and intensive care unit stay (p = .001; median difference 100.0 h; 95% CI 48-311) differed between patients with transient thrombosis and those with thrombosis lasting for more than 3 days. CONCLUSION: Internal jugular vein thrombosis was frequently detected by ultrasound following central venous catheterization in pediatric surgical patients. Multiple insertion attempts may be associated with the incidence of thrombosis. The clinical relevance of thrombi detected via ultrasound surveillance has not been determined.

Author Correction: Transcription Factor HOXA9 is Linked to the Calcification and Invasion of Papillary Thyroid Carcinoma.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Seroconversion of red blood cell antibody in ABO-incompatible living donor liver transplantation -a case report.

BACKGROUND: Liver transplantation usually requires blood transfusion, and a red blood cell (RBC) antibody screen is essential for the prevention of a hemolytic reaction. Since proper ABO-compatible grafts are lacking, ABO-incompatible living donor liver transplantation (ABO-i LDLT) with desensitization is a feasible therapy. Desensitization includes intravenous rituximab injection and plasmapheresis before surgery. CASE: A 60-year-old female was diagnosed with hepatitis B virus-related hepatocellular carcinoma and planned for ABO-i LDLT. She tested positive in a RBC antibody screen over two years; however, she tested negative for the test after desensitization. Clinicians noted the seroconversion during induction, and thus, a delay in the preparation of adequate packed RBC was unavoidable. CONCLUSIONS: Even when the latest RBC antibody screen is negative after immunosuppression, clinicians should consider the possibility of a prior positive result to promote safer medical treatment and management.

Transcription Factor HOXA9 is Linked to the Calcification and Invasion of Papillary Thyroid Carcinoma.

Calcification is important for the diagnosis of papillary thyroid carcinoma (PTC). Runt-related transcription factor 2 (RUNX2), a master transcription factor associated with osteogenic differentiation, is reportedly related to PTC calcification and invasiveness. However, its regulatory role in this process is somewhat uncharacterized. Here, we attempted to identify genes that regulate RUNX2 and clarify its function in PTC carcinogenesis and calcification. The expression of RUNX2-upstream genes was evaluated by real-time PCR in Nthy-Ori 3-1 normal thyroid cells and TPC1 and BHP10-3 PTC cell lines. Luciferase and chromatin immunoprecipitation assays were performed with candidate genes after cloning the RUNX2 promoter. We found that RUNX2 promoter activity was enhanced by homeobox family A9 (HOXA9). Over-expression of HOXA9 was found to enhance alkaline phosphatase activity, mineralization, and in vitro tumour cell migration and invasion, whereas downregulation had the opposite effects. These results indicate that HOXA9, a positive regulator of RUNX2, can enhance calcification, migration, and invasion in PTC. Our data improve the understanding of the molecular mechanisms of microcalcification in PTC as well as tumorigenesis.

STRA6 as a possible candidate gene for pathogenesis of osteoporosis from RNA­seq analysis of human mesenchymal stem cells.

To identify novel candidate genes associated with osteoporosis, RNA­sequence analysis of human mesenchymal stem cells (hMSCs) from patients with osteoporosis (G3) and osteopenia (G2), and healthy controls (G1) was performed. Differentially expressed genes (DEGs) from among the three groups were identified. DEGs were separated into nine groups according to their gene expression patterns: UU (up and up), UF (up and flat), UD (up and down), FU (flat and up), FF (flat and flat), FD (flat and down), DU (down and up), DF (down and flat), and DD (down and down). Among the 42 DEGs between G3 and G1, eight candidate genes, namely stimulated by retinoic acid 6 (STRA6), melanophilin, neurotrophic receptor tyrosine kinase 2, cartilage oligomeric matrix protein, collagen type XI α 1 chain, integrin subunit ß 2, monooxygenase DBH­like 1 and selenoprotein P, were selected, as they demonstrated consistent gene expression patterns of UU, FU, FD, and DD. Among these eight genes, STRA6 was highly expressed in the osteoporosis group and based on additional data from quantitative polymerase chain reaction analysis, it was selected for further study. In order to investigate whether STRA6 served a functional role in osteoblast or adipocyte differentiation, the effects of STRA6 expression changes in pluripotent stem cell C3H10T1/2, preosteoblast MC3T3­E1 and stromal ST2 cell lines were examined. Bone morphogenetic protein 2 enhanced STRA6 expression only at the early stage of osteoblast differe-ntiation, and overexpression of STRA6 temporally inhibited the expression of osteoblastogenesis markers, including runt related transcription factor 2, bone sialoprotein and osteocalcin. Furthermore, the knockdown of STRA6 slightly enhanced nodule formation at the late stage of osteoblast differentiation, and overexpression of STRA6 in ST2 cells enhanced adipocyte differentiation. Taken together, STRA6 expression could be associated with the pathogenesis of osteoporosis by promoting adipocyte differentiation over osteoblast differentiation in the hMSC population.

Transcriptional profiling of human femoral mesenchymal stem cells in osteoporosis and its association with adipogenesis.

Genetic alterations are major contributing factors in the development of osteoporosis. Osteoblasts and adipocytes share a common origin, mesenchymal stem cells (MSCs), and their genetic determinants might be important in the relationship between osteoporosis and obesity. In the present study, we aimed to isolate differentially expressed genes (DEGs) in osteoporosis and normal controls using human MSCs, and elucidate the common pathways and genes related to osteoporosis and adipogenesis. Human MSCs were obtained from the bone marrow of femurs from postmenopausal women during orthopedic surgeries. RNA sequencing (RNA-seq) was carried out using next-generation sequencing (NGS) technology. DEGs were identified using RNA-seq data. Ingenuity pathway analysis (IPA) was used to elucidate the common pathway related to osteoporosis and adipogenesis. Candidate genes for the common pathway were validated with other independent osteoporosis and obese subjects using RT-PCR (reverse transcription-polymerase chain reaction) analysis. Fifty-three DEGs were identified between postmenopausal osteoporosis patients and normal bone mineral density (BMD) controls. Most of the genetic changes were related to the differentiation of cells. The nuclear receptor subfamily 4 group A (NR4A) family was identified as possible common genes related to osteogenesis and adipogenesis. The expression level of the mRNA of NR4A1 was significantly higher in osteoporosis patients than in controls (p=0.018). The expression level of the mRNA of NR4A2 was significantly higher in obese patients than in controls (p=0.041). Some genetic changes in MSCs are involved in the pathophysiology of osteoporosis. The NR4A family might comprise common genes related to osteoporosis and obesity.

GATA4 negatively regulates bone sialoprotein expression in osteoblasts.

GATA4 has been reported to act as a negative regulator in osteoblast differentiation by inhibiting the Dlx5 transactivation of Runx2 via the attenuation of the binding ability of Dlx5 to the Runx2 promoter region. Here, we determine the role of GATA4 in the regulation of bone sialoprotein (Bsp) in osteoblasts. We observed that the overexpression of Runx2 or Sox9 induced the Bsp expression in osteoblastic cells. Silencing GATA4 further enhanced the Runx2- and Sox9-mediated Bsp promoter activity, whereas GATA4 overexpression down-regulated Bsp promoter activity mediated by Runx2 and Sox9. GATA4 also interacted with Runx2 and Sox9, by attenuating the binding ability of Runx2 and Sox9 to the Bsp promoter region. Our data suggest that GATA4 acts as a negative regulator of Bsp expression in osteoblasts. [BMB Reports 2016; 49(6): 343-348].

GATA4 negatively regulates osteoblast differentiation by downregulation of Runx2.

Osteoblasts are specialized mesenchymal cells that are responsible for bone formation. In this study, we examine the role of GATA4 in osteoblast differentiation. GATA4 was abundantly expressed in preosteoblast cells and gradually down-regulated during osteoblast differentiation. Overexpression of GATA4 in osteoblastic cells inhibited alkaline phosphatase activity and nodule formation in osteogenic conditioned cell culture system. In addition, overexpression of GATA4 attenuated expression of osteogenic marker genes, including Runx2, alkaline phosphatase, bone sialoprotein, and osteocalcin, all of which are important for osteoblast differentiation and function. Overexpression of GATA4 attenuated Runx2 promoter activity, whereas silencing of GATA4 increased Runx2 induction. We found that GATA4 interacted with Dlx5 and subsequently decreased Dlx5 binding activity to Runx2 promoter region. Our data suggest that GATA4 acts as a negative regulator in osteoblast differentiation by downregulation of Runx2.

Assessment of lifestyle effects on the levels of free oxygen radicals in the korean population.

BACKGROUND: As many studies revealed that oxidative stress due to the imbalance of reactive oxygen species (ROS) and antioxidant capacity is related with pathologic processes such as cardiovascular diseases, diabetes, as well as aging and obesity, the relationship between lifestyle and oxidative stress has recently gained much medical attention. However, little information exists on the effects of lifestyle on ROS in Korea. In this study, we investigated the effects of lifestyle on free oxygen radical levels in men and women in Korea. METHODS: A total of 138 adults participated in this study from September 2007 to June 2010 at a health promotion center and department of family medicine. Information on the lifestyle of each participant was obtained by questionnaire. Biochemical markers and a free oxygen radical test (FORT) were also measured. RESULTS: The average age was 47.28 ± 10.85 years and 79.7% were male. High sensitivity C-reactive protein (hs-CRP; r = 0.418, P = 0.012), triglycerides (r = -0.243, P = 0.008), hemoglobin (r = -0.445, P < 0.001), total protein (r = 0.210, P = 0.036), creatinine (r = -0.294, P = 0.001), fruit intake per day (P = 0.047), and smoking (P = 0.003) were related to the FORT levels in univariate analysis. Multiple linear regression analysis showed that hs-CRP (P = 0.039) was an independent predictor of serum FORT values. This statistical model can explain 78% of the variance in FORT values. CONCLUSION: This result suggests that hs-CRP showed a statistically significant positive association with FORT values. Further studies on the relationship between lifestyle and antioxidant capacity as well as ROS seem to be warranted to evaluate the overall effect of oxidative stress.

Negative feedback control of osteoclast formation through ubiquitin-mediated down-regulation of NFATc1.

The regulation of NFATc1 expression is important for osteoclast differentiation and function. Herein, we demonstrate that macrophage-colony-stimulating factor induces NFATc1 degradation via Cbl proteins in a Src kinase-dependent manner. NFATc1 proteins are ubiquitinated and rapidly degraded during late stage osteoclastogenesis, and this degradation is mediated by Cbl-b and c-Cbl ubiquitin ligases in a Src-dependent manner. In addition, NFATc1 interacts endogenously with c-Src, c-Cbl, and Cbl-b in osteoclasts. Overexpression of c-Src induces down-regulation of NFATc1, and depletion of Cbl proteins blocks NFATc1 degradation during late stage osteoclastogenesis. Taken together, our data provide a negative regulatory mechanism by which macrophage-colony-stimulating factor activates Src family kinases and Cbl proteins, and subsequently, induces NFATc1 degradation during osteoclast differentiation.

Silibinin inhibits osteoclast differentiation mediated by TNF family members.

Silibinin is a polyphenolic flavonoid compound isolated from milk thistle (Silybum marianum), with known hepatoprotective, anticarcinogenic, and antioxidant effects. Herein, we show that silibinin inhibits receptor activator of NF-κB ligand (RANKL)-induced osteoclastogenesis from RAW264.7 cells as well as from bone marrow-derived monocyte/macrophage cells in a dose-dependent manner. Silibinin has no effect on the expression of RANKL or the soluble RANKL decoy receptor osteoprotegerin (OPG) in osteoblasts. However, we demonstrate that silibinin can block the activation of NF-κB, c-Jun N-terminal kinase (JNK), p38 mitogen-activated protein (MAP) kinase, and extracellular signal-regulated kinase (ERK) in osteoclast precursors in response to RANKL. Furthermore, silibinin attenuates the induction of nuclear factor of activated T cells (NFAT) c1 and osteoclast-associated receptor (OSCAR) expression during RANKL-induced osteoclastogenesis. We demonstrate that silibinin can inhibit TNF-α-induced osteoclastogenesis as well as the expression of NFATc1 and OSCAR. Taken together, our results indicate that silibinin has the potential to inhibit osteoclast formation by attenuating the downstream signaling cascades associated with RANKL and TNF-α.

Regulatory mechanism of NFATc1 in RANKL-induced osteoclast activation.

NFATc1 is a master regulator of RANKL-induced osteoclast differentiation and herein we investigate the regulatory mechanism of NFATc1 in osteoclast activation. Inactivation of NFATc1 strongly attenuates RANKL-induced bone resorption and overexpression of a constitutively active form of NFATc1 in osteoclasts induces formation of actin rings and resorption pits on dentin slices. We demonstrate that NFATc1 binds directly to the promoter regions of its target genes and induces expression of various genes, including LTBP3, ClC7, cathepsin K, MMP9, and c-Src, which are key players in bone resorption. Thus, NFATc1 is essential for RANKL-induced osteoclast activation via up-regulation of osteoclast-activating genes.

The mechanism of osteoclast differentiation induced by IL-1.

IL-1 is a potent cytokine that can induce bone erosion in inflammatory sites such as rheumatoid joint regions via activation of osteoclasts. Not only is IL-1 capable of activating osteoclasts, but it is also a key cytokine involved in the differentiation, multinucleation, and survival of osteoclasts. Herein, we show that IL-1 has the potential to drive osteoclast differentiation via a receptor activator of NF-kappaB ligand (RANKL)/RANK-independent mechanism. Although IL-1 has a synergistic effect on RANKL-induced osteoclast formation, IL-1 alone cannot induce osteoclast differentiation from osteoclast precursors (bone marrow-derived macrophages (BMMs)) due to a lack of IL-1 signaling potential in these cells. However, we demonstrate that overexpression of the IL-1RI receptor in BMMs or induction of IL-1RI by c-Fos overexpression enables IL-1 alone to induce the formation of authentic osteoclasts by a RANKL/RANK-independent mechanism. The expression of IL-1RI is up-regulated by RANKL via c-Fos and NFATc1. Furthermore, the addition of IL-1 to IL-1RI overexpressing BMMs (IL-1/IL-1RI) strongly activates NF-kappaB, JNK, p38, and ERK which is a hallmark gene activation profile of osteoclastogenesis. Interestingly, IL-1/IL-1RI does not induce expression of c-Fos or NFATc1 during osteoclast differentiation, although basal levels of c-Fos and NFATc1 seem to be required. Rather, IL-1/IL-1RI strongly activates MITF, which subsequently induces osteoclast-specific genes such as osteoclast-associated receptor and tartrate-resistant acid phosphatase. Together, these results reveal that IL-1 has the potential to induce osteoclast differentiation via activation of microphthalmia transcription factor under specific microenvironmental conditions.

PCPH/ENTPD5 expression confers to prostate cancer cells resistance against cisplatin-induced apoptosis through protein kinase Calpha-mediated Bcl-2 stabilization.

Prostate cancer (PCa) frequently develops antiapoptotic mechanisms and acquires resistance to anticancer drugs. Therefore, identifying PCa drug resistance determinants should facilitate designing more effective chemotherapeutic regimens. Recently, we described that the PCPH protein becomes highly expressed in human prostatic intraepithelial neoplasia and in PCa, and that the functional interaction between PCPH and protein kinase Cdelta (PKCdelta) increases the invasiveness of human PCa. Here, we report that the functional interaction between PCPH and a different PKC isoform, PKCalpha, confers resistance against cisplatin-induced apoptosis to PCa cells. This interaction elicits a mechanism ultimately resulting in the posttranslational stabilization and subsequent elevated expression of Bcl-2. Stable knockdown of either PCPH, mt-PCPH, or PKCalpha in PCa cells decreased Ser70-phosphorylated Bcl-2 and total Bcl-2 protein, thereby increasing their cisplatin sensitivity. Conversely, forced expression of the PCPH protein or, in particular, of the mt-PCPH oncoprotein increased the levels of phosphorylated PKCalpha concurrently with those of Ser70-phosphorylated and total Bcl-2 protein, thus promoting cisplatin resistance. Consistently, Bcl-2 knockdown sensitized PCa cells to cisplatin treatment and, more importantly, reversed the cisplatin resistance of PCa cells expressing the mt-PCPH oncoprotein. Moreover, reexpression of Bcl-2 in PCPH/mt-PCPH knockdown PCa cells reversed the cisplatin sensitization caused by PCPH or mt-PCPH down-regulation. These findings identify PCPH and mt-PCPH as important participants in the chemotherapy response of PCa cells, establish a role for PCPH-PKCalpha-Bcl-2 functional interactions in the drug response process, and imply that targeting PCPH expression before, or simultaneously with, chemotherapy may improve the treatment outcome for PCa patients.

Upstream stimulatory factors regulate OSCAR gene expression in RANKL-mediated osteoclast differentiation.

Receptor activator of nuclear factor kappaB ligand (RANKL) induces osteoclast differentiation from hematopoietic precursors via regulation of various transcription factors. Here, we show that upstream stimulatory factors (USFs), which are basic helix-loop-helix leucine zipper transcription factors, are involved in RANKL-mediated osteoclastogenesis and regulation of osteoclast-associated receptor (OSCAR) gene expression, which is an important costimulatory receptor for osteoclast differentiation. USF-1 and USF-2 are expressed in hematopoietic osteoclast precursors and mature osteoclasts. Interestingly, down-regulation of USF-1 by RNA interference or overexpression of a dominant-negative form of USF-1 attenuates osteoclast formation and expression of OSCAR during osteoclastogenesis. Promoter analysis and chromatin immunoprecipitation assays reveal that USFs bind directly to an E-box site in the OSCAR promoter region and activate OSCAR. In addition, USFs interact and cooperate with nuclear factor of activated T cells c1 in regulating OSCAR gene expression. Taken together, our results indicate that USFs serve as modulators in the induction of OSCAR and RANKL-mediated osteoclastogenesis.