Bariatric surgery and weight loss in the short- and long-term: Evidence from NHANES 2015-2018.

The present study aimed to examine short- and long-term weight change in a nationally representative sample of US adults who reportedly underwent bariatric surgery. Individuals aged 20-64 at survey from the US National Health and Nutrition Examination Survey 2015-2018 were included in the analyses (n = 6776). The primary comparison groups include 62 participants who underwent bariatric surgery, 1531 eligible but did not receive surgery, and 5183 not eligible for bariatric surgery. After adjusting for demographic characteristics and comorbidity, adults who reported receiving bariatric surgery were 5.0 times (4.0-6.0) more likely to achieve at least 20% weight loss from maximum weight relative to those who were eligible but reported no surgery. The likelihood appeared to be higher when surgery was performed within 10 years (short-term, PR 5.5, 95% CI: 4.0, 7.0) relative to surgeries that were performed for 10 or more years (long-term, PR 3.6, 95% CI: 2.0, 5.3). In this nationally representative sample of US adults, respondents who received bariatric surgery achieved substantial and significant weight loss compared with those who were eligible and did not receive bariatric surgery. Weight loss appeared to be most apparent in the short term and persisted over the long term.

Outcomes associated with the use of a new powered circular stapler for left-sided colorectal reconstructions: a propensity score matching-adjusted indirect comparison with manual circular staplers.

BACKGROUND: This was a retrospective, matching-adjusted indirect comparison of clinical outcomes between patients from a single-arm trial of the ECHELON CIRCULAR™ Powered Stapler (ECP) and those from a historical cohort of patients who underwent left-sided colorectal resection using conventional manual circular staplers, extracted from the Premier Healthcare Database. METHODS: Patients in the ECP trial cohort were propensity score matched to those in the historical cohort through nearest neighbor matching. Outcomes included 30-day readmission rates; length of stay (LOS) for the index admission; rates of anastomotic leak, pelvic abscess, ileus/small bowel obstruction, infection, bleeding, and stoma creation. RESULTS: The study included 168 patients in the ECP trial cohort and 4544 patients in the historical cohort; 165 ECP trial patients were matched to 1348 historical cohort patients. After matching, conversions were more prevalent in the historical cohort than the ECP trial cohort (4.2% ECP vs. 10.2% historical, p = 0.001). Relative to the historical cohort, the ECP trial cohort had statistically significant lower rates of 30-day inpatient readmission (6.1% vs. 10.8%, p = 0.019), anastomotic leak (1.8% vs. 6.9%, p < 0.001), ileus/small bowel obstruction (4.8% vs. 14.7%, p < 0.001), infection (1.8% vs. 5.7%, p = 0.001), and bleeding (1.8% vs. 9.2%, p < 0.001) during the index admission or within 30 days thereafter. No statistically significant differences in rates of pelvic abscess, stoma creation, or LOS were found between the two cohorts. Three sensitivity analyses to address the difference in conversion rates yielded largely consistent results, with loss of statistical significance for inpatient admission in some cases. This study is limited by its potential for differences in unmeasurable factors between the ECP trial and historical cohorts. CONCLUSIONS: In this study, the ECP trial cohort had lower incidence proportions of several surgical complications as compared with the historical cohort. Further controlled prospective clinical studies are needed to confirm the validity of this finding.

Safety and effectiveness of REALIZE adjustable gastric band: 5-year prospective study.

BACKGROUND: The long-term safety results of the REALIZE (Ethicon Endo-Surgery, Inc., Cincinnati, OH) adjustable gastric band collected in this prospective, multicenter study in patients with morbid obesity are presented. OBJECTIVES: To determine the reoperation rate, including band revisions, replacements, and explants, resulting from a serious adverse device-related event through years 4 and 5. Various efficacy measures were also assessed as secondary objectives. SETTING: Nine academic and/or private institutions. METHODS: The participating institutions enrolled 303 patients, who were then assessed on an annual basis, with 231 patients completing 5 years of follow-up. The study parameters included reoperation rates, changes in percentage of excess weight loss (%EWL), and changes in body mass index (BMI), as well as parameters of diabetes and dyslipidemia. Quality of life was assessed using the Short Form (SF)-36 and the Impact of Weight on Quality of Life-Lite questionnaires. RESULTS: The reoperation rate due to a serious adverse event in this population at 5 years after implantation with the REALIZE gastric band was 8.9%. The most common serious adverse event was band slippage, which affected 6.9% of the study population. The mean %EWL was 35.6% ± 26.84%, and the decrease in mean BMI was -7.01 ± 5.45 kg/m2 at 5 years. Patients experienced improvements in mean glycated hemoglobin and serum lipid levels, in addition to improvements in the quality of life measures. CONCLUSION: No new safety concerns were identified during the 5 years of follow-up. Although the results of this study did not meet the predefined safety criteria of 8% or less, the safety profile and long-term effectiveness observed in this study are consistent with those in the current literature.

Obesity Progression Between Young Adulthood and Midlife and Incident Arthritis: A Retrospective Cohort Study of US Adults.

OBJECTIVE: To examine the association between weight change from young adulthood to midlife and the risk of incident arthritis. METHODS: Using data from the National Health and Nutrition Examination Survey, we categorized participants into weight-change categories based on their recalled weight during young adulthood and midlife. We estimated the association of weight change and developing an arthritis condition over 10 years using adjusted Cox models. Findings were extrapolated to the US population to determine the proportion of incident arthritis cases that could be averted if the entire population maintained a normal body mass index (BMI) in young adulthood and midlife. RESULTS: Among our sample of adults who were ages 40-69 years at their midlife weight measure (n = 13,669), 3,603 developed an arthritis condition. Compared with adults who maintained a normal-normal BMI, the normal-overweight, normal-obese, overweight-obese, and obese-obese groups had a significantly elevated risk of incident arthritis conditions. The obese-overweight group had a lower risk of incident arthritis conditions compared with the obese-obese group and a comparable risk to the overweight-overweight group. Nearly one-fourth of incident arthritis cases, corresponding to 2.7 million individuals, would have been averted under the hypothetical scenario where all individuals maintained normal weight from young adulthood to midlife. CONCLUSION: Weight loss from young adulthood to midlife was associated with a substantially reduced risk of developing an arthritis condition. We found no evidence of residual risk from having been heavier earlier in life. Our findings highlight the critical need to expand obesity treatment and prevention to achieve meaningful reductions in the burden of arthritis.

Assessment of a circular powered stapler for creation of anastomosis in left-sided colorectal surgery: A prospective cohort study.

BACKGROUND: Circular staplers perform a critical function for creation of anastomoses in colorectal surgeries. Powered stapling systems allow for reduced force required by surgeons to fire the device and may provide advantages for creating a secure anastomosis. The objective of this study was to evaluate the clinical performance of a novel circular powered stapler in a post-market setting, during left-sided colectomy procedures. MATERIALS AND METHODS: Consecutive subjects underwent left-sided colorectal resections that included anastomosis performed with the ECHELON CIRCULAR™ Powered Stapler (ECP). The primary endpoint was the frequency in which a stapler performance issue was observed. Secondary endpoints included evaluation of ease of use of the device via a surgeon satisfaction questionnaire, and monitoring/recording of procedure-related adverse events (AEs). RESULTS: A total of 168 anastomoses were performed with the ECP. Surgical approaches included robotic-assisted (n = 74, 44.0%), laparoscopic (n = 71, 42.3%), open (n = 20, 11.9%), and hand-assisted minimally invasive (n = 3, 1.8%) procedures. There were 22 occurrences of device performance issues in 20 (11.9%) subjects during surgery. No positive intraoperative leak tests were observed, and only 1 issue was related to a procedure-related AE or surgical complication, which was an instance of incomplete surgical donut necessitating re-anastomosis. Postoperative anastomotic leaks were experienced in 4 (2.4%) subjects. Clavien-Dindo classification of all AEs indicated that 92.0% were Grades I or II. Participating surgeons rated the ECP as easier to use compared to previously used manual circular staplers in 85.7% of procedures. CONCLUSION: The circular powered stapler exhibited few clinically relevant performance issues, an overall favorable safety profile, and ease of use for creation of left-sided colon anastomoses.

Association of Weight Loss Between Early Adulthood and Midlife With All-Cause Mortality Risk in the US.

Importance: Describing potential mortality risk reduction associated with weight loss between early adulthood and midlife is important for informing primary and secondary prevention efforts for obesity. Objective: To examine the risk of all-cause mortality among adults who lost weight between early adulthood and midlife compared with adults who were persistently obese over the same period. Design, Setting, and Participants: Combined repeated cross-sectional analysis was conducted using data from the National Health and Nutrition Examination Survey III (1988-1994) and continuous waves collected in 2-year cycles between 1999 and 2014. The data analysis was conducted from February 10, 2019, to April 20, 2020. Individuals aged 40 to 74 years at the time of survey (baseline) were included in the analyses (n = 24 205). Exposures: Weight history was assessed by self-reported weight at age 25 years, at 10 years before baseline (midlife: mean age, 44 years; interquartile range, 37-55), and measured weight at baseline. Body mass index (BMI) (calculated as weight in kilograms divided by height in meters squared) at each time was categorized as normal (18.5-24.9), overweight (25.0-29.9), and obese (≥30.0). Weight change patterns were assessed from age 25 years (early adulthood) to 10 years before baseline (midlife). Main Outcomes and Measures: Incident all-cause mortality using linked data from the National Death Index. Results: Of the 24 205 participants, 11 617 were women (49.0%) and 11 567 were non-Hispanic White (76.9%). The mean (SD) BMI was 29.0 (6.1) at baseline. During a mean (SD) follow-up of 10.7 (7.2) years, 5846 deaths occurred. Weight loss from obese to overweight was associated with a 54% (hazard ratio, 0.46; 95% CI, 0.27-0.77) reduction in mortality risk compared with individuals with stable obesity between early adulthood and midlife. An estimated 3.2% (95% CI, 1.6%-4.9%) of early deaths could have been avoided if those who maintained an obese BMI instead lost weight to an overweight BMI by midlife. Overall, an estimated 12.4% (95% CI, 8.1%-16.5%) of early deaths may be attributable to having weight in excess of the normal BMI range at any point between early and mid-adulthood. Conclusions and Relevance: In this study, weight loss from obesity to overweight between early adulthood through midlife appeared to be associated with a mortality risk reduction compared with persistent obesity. These findings support the importance of population-based approaches to preventing weight gain across the life course and a need for greater emphasis on treating obesity early in life.

Evaluation of a Powered Vascular Stapler in Video-Assisted Thoracic Surgery Lobectomy.

BACKGROUND: A narrow-profile powered vascular stapler (PVS) was developed to provide superior access and precise staple placement in thoracic procedures. The objective of this study was to determine if the PVS would yield an equivalent rate of hemostatic interventions compared with standard of care (SOC) staplers in video-assisted thoracoscopic surgery lobectomy. MATERIALS AND METHODS: A randomized, controlled, multicenter study was conducted comparing PVS with SOC staplers in lobectomies performed for non-small cell lung cancer. The primary performance endpoint was the incidence of intraoperative hemostatic interventions, and the primary safety endpoint was the frequency of postoperative bleeding-related interventions. RESULTS: A total of 98 subjects participated in the SOC group and 103 in the PVS group. Rates of intraoperative hemostatic interventions were 5.3% and 8.3% for the SOC and PVS groups, respectively. These rates were not statistically different (P = 0.137), although the upper bound of the 95% confidence interval for the difference in intervention rates between PVC and SOC exceeded a predefined 3% criterion for equivalence. Simple compressions were performed more frequently in the PVS subjects, which accounted for the higher intervention rate in this group. Postoperative interventions for bleeding were required in one SOC subject (1.0%) and one subject from the PVS group (0.9%). Procedure-related adverse events occurred in 21 (21.9%) SOC subjects and 23 (21.9%) PVS subjects, with no adverse events related to use of the study devices. CONCLUSIONS: The PVS exhibited similar overall safety and effectiveness to SOC staplers in video-assisted thoracoscopic surgery lobectomy.

Clinical outcomes following an initial experience with a novel powered vascular stapler in video-assisted thoracoscopic lobectomies: results of a Chinese multi-center study.

BACKGROUND: The development of minimally invasive surgical approaches has revolutionized surgical care and greatly improved surgical outcomes. This study aimed to evaluate the clinical effectiveness of a powered vascular stapler (PVS) during video-assisted thoracoscopic surgery (VATS) lobectomy. METHODS: This prospective, multi-center, post-market clinical study in China enrolled 50 patients with either a suspected or formal diagnosis of clinical stage IA to IIB non-small cell lung cancer (NSCLC) scheduled for VATS lobectomy. The clinical effectiveness of the PVS for successful pulmonary artery (PA)/pulmonary vein (PV) transection was evaluated. In addition, the surgeon's stress, device usability, and surgeon satisfaction were measured using multiple questionnaires. RESULTS: A total of 167 PAs/PVs were transected with 3 (1.8%) requiring intra-operative intervention. Fourteen of the 50 patients (28.0%) experienced at least one adverse event (AE), among whom 5 (10.0%) suffered from serious AEs. There were no postoperative hemorrhagic complications related to transection of the PA/PV with PVS. Surgeon satisfaction was surveyed by questionnaire after each of the 50 procedures resulting in a 96% reported satisfaction with device usability, specifically related to a low stress load and an increase in work efficiency. CONCLUSIONS: For VATS lobectomy, the PVS demonstrated a positive surgical effectiveness and value in cognitive and physical distress reduction. Complications following VATS lobectomy to treat NSCLC were generally low and as expected. Intraoperative complications were few and there were no postoperative complications related to the transection of the PA and PV during VATS lobectomy. Favorable results were reported on the surgeon satisfaction questionnaire regarding usability and surgeon stress.

The contribution of obesity to prescription opioid use in the United States.

The prevalence of obesity has grown rapidly over the past several decades and has been accompanied by an increase in the prevalence of chronic pain and prescription opioid use. Obesity, through its association with pain, may represent an important contributor to opioid use. This cross-sectional study investigated the relationship between obesity and prescription opioid use among adults aged 35 to 79 years using data from the National Health and Nutrition Examination Survey (NHANES, 2003-2016). Relative to normal weight, body mass indices in the overweight {odds ratio (OR), 1.11 (confidence interval [CI], 0.88-1.39)}, obese I (OR, 1.26 [CI, 1.01-1.57]), obese II (OR, 1.69 [CI, 1.34-2.12]), and obese III (OR, 2.33 [CI, 1.76-3.08]) categories were associated with elevated odds of prescription opioid use. The association between excess weight and opioid use was stronger for chronic opioid use than for use with a duration of less than 90 days (P-value, <0.001). We estimated that 14% (CI, 9%-19%) of prescription opioid use at the population level was attributable to obesity, suggesting there might have been 1.5 million fewer opioid users per year under the hypothetical scenario where obese individuals were instead nonobese (CI, 0.9-2.0 million users). Back pain, joint pain, and muscle/nerve pain accounted for the largest differences in self-reported reasons for prescription opioid use across obesity status. Although interpretation is limited by the cross-sectional nature of the associations, our findings suggest that the obesity epidemic may be partially responsible for the high prevalence of prescription opioid use in the United States.

A novel approach to glycemic control in type 2 diabetes mellitus, partial jejunal diversion: pre-clinical to clinical pathway.

OBJECTIVE: To explore partial jejunal diversion (PJD) via a side-to-side jejuno-jejunostomy for improved glycemic control in type 2 diabetes mellitus (T2DM). PJD is an anatomy-sparing, technically simple surgery in comparison to the predominate metabolic procedures, Roux-en-Y gastric bypass (RYGB) and sleeve gastrectomy (SG). Positive results in a rodent model prompted a human proof-of-concept study. RESEARCH DESIGN AND METHODS: Pre-clinically, 71 rats were studied in a model of metabolic dysfunction induced by a high-fat diet; 33 animals undergoing one of two lengths of PJD were compared with 18 undergoing sham, 10 RYGB and 10 jejuno-ileal bypass. Clinically, 15 adult subjects with treated but inadequately controlled T2DM (hemoglobin A1c (HbA1c) of 8.0%-11.0%), body mass index of 27.0-40.0 kg/m2, and C peptide ≥3 ng/mL were studied. Follow-up was at 2 weeks, and 3, 6, 9, and 12 months post-PJD. RESULTS: Pre-clinically, positive impacts with PJD on glucose homeostasis, cholesterol, and body composition versus sham control were demonstrated. Clinically, PJD was performed successfully without serious complications. Twelve months post-surgery, the mean (SD) reduction from baseline in HbA1c was 2.3% (1.3) (p<0.01). CONCLUSIONS: PJD may provide an anatomy sparing, low-risk, intervention for poorly controlled T2DM without significant alteration of the patient's lifestyle. The proof-of-concept study is limited by a small sample size and advanced disease, with 80% of participants on insulin and a mean time since diagnosis of over 10 years. Further study is warranted. TRIAL REGISTRATION NUMBER: NCT02283632; Pre-results.

A randomized study comparing outcomes of stapled and hand-sutured anastomoses in patients undergoing open gastrointestinal surgery.

Although stapling is an alternative to hand-suturing in gastrointestinal surgery, recent trials specifically designed to evaluate differences between the two in surgery time, anastomosis time, and return to bowel activity are lacking. This trial compared the outcomes of the two in subjects undergoing open gastrointestinal surgery. Adult subjects undergoing emergency or elective surgery requiring a single gastric, small, or large bowel anastomosis were enrolled into this open-label, prospective, randomized, interventional, parallel, multicenter, controlled trial. Randomization was assigned in a 1:1 ratio between the hand-sutured group (n = 138) and the stapled group (n = 142). Anastomosis time, surgery time, and time to bowel activity were collected and compared as primary endpoints. A total of 280 subjects were enrolled from April 2009 to September 2010. Only the time of anastomosis was significantly different between the two arms: 17.6 ± 1.90 min (stapled) and 20.6 ± 1.90 min (hand-sutured). This difference was deemed not clinically or economically meaningful. Safety outcomes and other secondary endpoints were similar between the two arms. Mechanical stapling is faster than hand-suturing for the construction of gastrointestinal anastomoses. Apart from this, stapling and hand-suturing are similar with respect to the outcomes measured in this trial.

The human phospholamban Arg14-deletion mutant localizes to plasma membrane and interacts with the Na/K-ATPase.

Depressed Ca-handling in cardiomyocytes is frequently attributed to impaired sarcoplasmic reticulum (SR) function in human and experimental heart failure. Phospholamban (PLN) is a key regulator of SR and cardiac function, and PLN mutations in humans have been associated with dilated cardiomyopathy (DCM). We previously reported the deletion of the highly conserved amino acid residue arginine 14 (nucleic acids 39, 40 and 41) in DCM patients. This basic amino acid is important in maintaining the upstream consensus sequence for PKA phosphorylation of Ser 16 in PLN. To assess the function of this mutant PLN, we introduced the PLN-R14Del in cardiac myocytes of the PLN null mouse. Transgenic lines expressing mutant PLN-R14Del at similar protein levels to wild types exhibited no inhibition of the initial rates of oxalate-facilitated SR Ca uptake compared to PLN-knockouts (PLN-KO). The contractile parameters and Ca-kinetics also remained highly stimulated in PLN-R14Del cardiomyocytes, similar to PLN-KO, and isoproterenol did not further stimulate these hyper-contractile basal parameters. Consistent with the lack of inhibition on SR Ca-transport and contractility, confocal microscopy indicated that the PLN-R14Del failed to co-localize with SERCA2a. Moreover, PLN-R14Del did not co-immunoprecipitate with SERCA2a (as did WT-PLN), but rather co-immunoprecipitated with the sarcolemmal Na/K-ATPase (NKA) and stimulated NKA activity. In addition, studies in HEK cells indicated significant fluorescence resonance energy transfer between PLN-R14Del-YFP and NKAα1-CFP, but not with the NKA regulator phospholemman. Despite the enhanced cardiac function in PLN-R14Del hearts (as in PLN-knockouts), there was cardiac hypertrophy (unlike PLN-KO) coupled with activation of Akt and the MAPK pathways. Thus, human PLN-R14Del is misrouted to the sarcolemma, in the absence of endogenous PLN, and alters NKA activity, leading to cardiac remodeling.

The anti-apoptotic protein HAX-1 is a regulator of cardiac function.

The HS-1 associated protein X-1 (HAX-1) is a ubiquitously expressed protein that protects cardiomyocytes from programmed cell death. Here we identify HAX-1 as a regulator of contractility and calcium cycling in the heart. HAX-1 overexpression reduced sarcoplasmic reticulum Ca-ATPase (SERCA2) pump activity in isolated cardiomyocytes and in vivo, leading to depressed myocyte calcium kinetics and mechanics. Conversely, downregulation of HAX-1 enhanced calcium cycling and contractility. The inhibitory effects of HAX-1 were abolished upon phosphorylation of phospholamban, which plays a fundamental role in controlling basal contractility and constitutes a key downstream effector of the beta-adrenergic signaling cascade. Mechanistically, HAX-1 promoted formation of phospholamban monomers, the active/inhibitory units of the calcium pump. Indeed, ablation of PLN rescued HAX-1 inhibition of contractility in vivo. Thus, HAX-1 represents a regulatory mechanism in cardiac calcium cycling and its responses to sympathetic stimulation, implicating its importance in calcium homeostasis and cell survival.

Phospholamban overexpression in rabbit ventricular myocytes does not alter sarcoplasmic reticulum Ca transport.

Phospholamban has been suggested to be a key regulator of cardiac sarcoplasmic reticulum (SR) Ca cycling and contractility and a potential therapeutic target in restoring the depressed Ca cycling in failing hearts. Our understanding of the function of phospholamban stems primarily from studies in genetically altered mouse models. To evaluate the significance of this protein in larger mammalian species, which exhibit Ca cycling properties similar to humans, we overexpressed phospholamban in adult rabbit cardiomyocytes. Adenoviral-mediated gene transfer, at high multiplicities of infection, resulted in an insignificant 1.22-fold overexpression of phospholamban. There were no effects on twitch Ca-transient amplitude or decay under basal or isoproterenol-stimulated conditions. Furthermore, the SR Ca load and Na/Ca exchanger function were not altered. These apparent differences between phospholamban overexpression in rabbit compared with previous findings in the mouse may be due to a significantly higher (1.5-fold) endogenous phospholamban-to-sarco(endo)plasmic reticulum Ca-ATPase (SERCA) 2a ratio and potential functional saturation of SERCA2a by phospholamban in rabbit cardiomyocytes. The findings suggest that important species-dependent differences in phospholamban regulation of SERCA2a occur. In larger mammals, a higher fraction of SERCA2a pumps are regulated by phospholamban, and this may influence therapeutic strategies to enhance cardiac contractility and functional cardiac reserve.

Phospholamban overexpression in transgenic rabbits.

There has been considerable interest in pursuing phospholamban as a putative therapeutic target for overcoming depressed calcium handling in human heart failure. Studies predominantly done in mice have shown that phospholamban is a key regulator of sarcoplasmic reticulum calcium cycling and cardiac function. However, mice differ significantly from humans in how they regulate calcium, whereas rabbits better recapitulate human cardiac function and calcium handling. To investigate phospholamban's role in the rabbit heart, transgenic rabbits that overexpressed wild-type phospholamban in the ventricular cardiomyocytes and slow-twitch skeletal muscles were generated. Rabbits expressing high levels of phospholamban were not viable due to severe skeletal muscle wasting, the onset of cardiac pathology and early death. A viable transgenic line exhibited a 30% increase in PLN protein levels in the heart. These animals showed isolated foci of cardiac pathology, but cardiac function as well as the response to beta-adrenergic stimulation were normal. SR-calcium uptake measurements showed that the transgenic hearts had the expected reduced affinity for calcium. The data show that phospholamban-overexpressing transgenic rabbits differ markedly in phenotype from analogous transgenic mice in that rabbits are quite sensitive to alterations in phospholamban levels. Exceeding a relatively narrow window of phospholamban expression results in significant morbidity and early death.

Overexpression of histidine-rich Ca-binding protein protects against ischemia/reperfusion-induced cardiac injury.

OBJECTIVE: The histidine-rich Ca-binding protein (HRC) is a Ca-storage protein in cardiac sarcoplasmic reticulum. Recent transgenic studies revealed that this protein inhibits the maximal rates of sarcoplasmic reticulum Ca-transport, leading to cardiac dysfunction. In view of the role of sarcoplasmic reticulum Ca-cycling in myocardial ischemia/reperfusion injury, we designed this study to gain further insight into the role of HRC during ischemia/reperfusion. METHODS AND RESULTS: The transgenic mouse model with cardiac-specific overexpression of HRC was utilized and cardiac contractile parameters were assessed before and after ischemia/reperfusion injury by Langendorff perfusion. After a 20-min stabilization period, the hearts were subjected to 40 min of global ischemia, followed by 60 min of reperfusion. We found that although transgenic (TG) hearts showed depressed cardiac function (25%) compared to wild types (WTs) at baseline, they exhibited better recovery of left ventricular developed pressure (86.6+/-2.6% in TGs vs. 58.3+/-4.0% in WTs of pre-ischemic values, P<0.05) and higher rates of contraction and relaxation after ischemia/reperfusion than WTs. This improvement was accompanied by smaller infarcts (23.1+/-1.7% in TGs vs. 41.1+/-2.5% in WTs of infarct region-to-risk region ratio, P<0.05) and lower creatine kinase release. Notably, the extent of apoptotic cell death was significantly attenuated, as evidenced by decreased DNA fragmentation, upregulation of the antiapoptotic protein Bcl-2, and downregulation of the active caspases (3, 9 and 12) following ischemia/reperfusion in TG hearts, compared with WTs. Extension of these studies to an in vivo model of 30-min myocardial ischemia, via coronary artery occlusion, followed by 24-h reperfusion, showed that the infarct region-to-risk region ratio was 9+/-0.9% in TGs, compared with 20.4+/-2.9% in WTs (P<0.05). CONCLUSIONS: Our findings suggest that increased cardiac HRC expression protects against ischemia/reperfusion injury in the heart, resulting in improved recovery of function and reduced infarction.

Sarcoplasmic reticulum calcium overloading in junctin deficiency enhances cardiac contractility but increases ventricular automaticity.

BACKGROUND: Abnormal sarcoplasmic reticulum calcium (Ca) cycling is increasingly recognized as an important mechanism for increased ventricular automaticity that leads to lethal ventricular arrhythmias. Previous studies have linked lethal familial arrhythmogenic disorders to mutations in the ryanodine receptor and calsequestrin genes, which interact with junctin and triadin to form a macromolecular Ca-signaling complex. The essential physiological effects of junctin and its potential regulatory roles in sarcoplasmic reticulum Ca cycling and Ca-dependent cardiac functions, such as myocyte contractility and automaticity, are unknown. METHODS AND RESULTS: The junctin gene was targeted in embryonic stem cells, and a junctin-deficient mouse was generated. Ablation of junctin was associated with enhanced cardiac function in vivo, and junctin-deficient cardiomyocytes exhibited increased contractile and Ca-cycling parameters. Short-term isoproterenol stimulation elicited arrhythmias, including premature ventricular contractions, atrioventricular heart block, and ventricular tachycardia. Long-term isoproterenol infusion also induced premature ventricular contractions and atrioventricular heart block in junctin-null mice. Further examination of the electrical activity revealed a significant increase in the occurrence of delayed afterdepolarizations. Consistently, 25% of the junctin-null mice died by 3 months of age with structurally normal hearts. CONCLUSIONS: Junctin is an essential regulator of sarcoplasmic reticulum Ca release and contractility in normal hearts. Ablation of junctin is associated with aberrant Ca homeostasis, which leads to fatal arrhythmias. Thus, normal intracellular Ca cycling relies on maintenance of junctin levels and an intricate balance among the components in the sarcoplasmic reticulum quaternary Ca-signaling complex.

Phospholamban inhibits Ca-ATPase conformational changes involving the E2 intermediate.

We have used steady-state fluorescence spectroscopy in combination with enzyme kinetic assays to test the hypothesis that phospholamban (PLB) stabilizes the Ca-ATPase in the E2 intermediate state. The cardiac muscle Ca-ATPase (SERCA2a) isoform was expressed either alone or coexpressed with PLB in High-Five insect cells and was isolated as insect cell microsomes. Fluorescence studies of the Ca-ATPase covalently labeled with the probe 5-(2-((iodoacetyl)amino)ethyl)aminonaphthalene-1-sulfonic acid showed that PLB decreased the amplitude of the Ca-ATPase E2 --> E1 conformational transition by 45 +/- 3% and shifted the [Ca2+] dependence of the transition to higher Ca2+ levels (DeltaKCa = 230 nM), similar to the effect of PLB on Ca-ATPase activity. Similarly, PLB decreased the amplitude of Ca-ATPase phosphorylation by inorganic phosphate (Pi) by 55 +/- 2% and decreased slightly the affinity for Pi (DeltaK0.5 = 70 microM). However, PLB did not affect the Ca2+-dependent inhibition of Ca-ATPase phosphorylation by Pi. Finally, PLB decreased Ca-ATPase sensitivity to vanadate, increasing the IC50 value by 300 nM. The results suggest that PLB binding to Ca-ATPase stabilizes the enzyme in a conformation distinct from E2, decreasing the number of enzymes in the E2 state capable of undergoing ligand-dependent conformational changes involving the Ca-free E2 intermediate. The inability of conformation-specific ligands to fully convert this E2-like state into E1 or E2 implies that these states are not in a simple equilibrium relationship.

Identification of a novel phosphorylation site in protein phosphatase inhibitor-1 as a negative regulator of cardiac function.

Human and experimental heart failure is characterized by increases in type-1 protein phosphatase activity, which may be partially attributed to inactivation of its endogenous regulator, protein phosphatase inhibitor-1. Inhibitor-1 represents a nodal integrator of two major second messenger pathways, adenosine 3',5'-cyclic monophosphate (cAMP) and calcium, which mediate its phosphorylation at threonine 35 and serine 67, respectively. Here, using recombinant inhibitor-1 wild-type and mutated proteins, we identified a novel phosphorylation site in inhibitor-1, threonine 75. This phosphoamino acid was phosphorylated in vitro by protein kinase Calpha independently and to the same extent as serine 67, the previous protein kinase Calpha-identified site. Generation of specific antibodies for the phosphorylated and dephosphorylated threonine 75 revealed that this site is phosphorylated in rat and dog hearts. Adenoviral-mediated expression of the constitutively phosphorylated threonine 75 inhibitor-1 in isolated myocytes was associated with specific stimulation of type-1 protein phosphatase activity and marked inhibition of the sarcoplasmic calcium pump affinity for calcium, resulting in depressed contractility. Thus, phosphorylation of inhibitor-1 at threonine 75 represents a new mechanism of cardiac contractility regulation, partially through the alteration of sarcoplasmic reticulum calcium transport activity.

Histidine-rich Ca binding protein: a regulator of sarcoplasmic reticulum calcium sequestration and cardiac function.

Defects in the pathways that regulate cardiac sarcoplasmic reticulum (SR) calcium (Ca) cycling represent prime targets for driving the deterioration of function and progression to heart failure. We hypothesized that the histidine-rich Ca binding protein (HRC) in the SR may be involved in SR Ca cycling and that alterations in HRC levels would result in abnormal cardiac Ca homeostasis. In order to test this hypothesis, we generated transgenic mice with cardiac overexpression (3-fold) of HRC. Increased cardiac HRC levels were associated with impaired SR Ca uptake rates (35%) and attenuated cardiomyocyte Ca transient decay (38%), without alterations in peak Ca transients or SR Ca load. The depressed SR Ca sequestration was associated with attenuated rate of Ca extrusion via Na-Ca exchange. Triadin protein expression levels and L-type Ca channel current density were increased, while the channel inactivation kinetics were not altered. Impaired SR Ca uptake and delayed Ca decline rates triggered hypertrophy and compromised the heart's responses to increased stress by either hemodynamic overload or the aging process. By 18 months of age, cardiac remodeling deteriorated to congestive heart failure in transgenic mice. Collectively, these data suggest that HRC may be an integral regulatory protein involved in cardiac muscle SR Ca uptake and Ca homeostasis.