Factors Contributing to Delays to Accessing Appendectomy in Low- and Middle-Income Countries: A Scoping Review.

BACKGROUND: Appendicitis is one of the most common emergency surgical conditions worldwide. Delays in accessing appendectomy can lead to complications. Evidence on these delays in low- and middle-income countries (LMICs) is lacking. The aim of this review was to identify and synthesise the available evidence on delays to accessing appendectomy in LMICs. METHODS: This scoping review followed the Preferred Reporting Items for Systematic Reviews and Meta-analysis Extension for Scoping Reviews framework. The delays and their interconnectivity in LMICs were synthesised and interpreted using the Three Delays framework. We reviewed Africa Wide EBSCOhost, PubMed-Medline, Scopus, Web of Science, African Journals Online (AJOL), and Bioline databases. RESULTS: Our search identified 21 893 studies, of which 78 were included in the final analysis. All of the studies were quantitative. Fifty per cent of the studies included all three types of delays. Delays in seeking care were influenced by a lack of awareness of appendicitis symptoms, and the use of self and alternative medication, which could be linked to delays in receiving care, and the barrier refusal of medical treatment due to fear. Financial concerns were a barrier observed throughout the care pathway. CONCLUSION: This review highlighted the need for additional studies on delays to accessing appendectomy in additional LMICs. Our review demonstrates that in LMICs, persons seeking appendectomy present late to health-care facilities due to several patient-related factors. After reaching a health-care facility, accessing appendectomy can further be delayed owing to a lack of adequate hospital resources.

Surgical training during the COVID-19 pandemic - a single institution's trainee survey.

BACKGROUND: Few studies have assessed the impact of COVID-19 on surgical training in low- and middle-income countries. The aim of this study was to survey the effect of the COVID-19 pandemic on postgraduate surgical training, research and registrar wellbeing in South Africa. METHODS: A cross-sectional study was conducted as an online survey from 5 October 2020 to 1 December 2020. The study population was registrars from all surgical disciplines at the Faculty of Medicine and Health Sciences of Stellenbosch University. The survey consisted of 26 multiple-choice and five open-ended qualitative questions on the impact of COVID-19 on physical and mental wellbeing, skills acquisition and postgraduate research. RESULTS: Of 98 surgical registrars, 35 (36%) responded. Twenty-three (65.7%) reported missed planned surgical rotations, 30 (85.7%) decreased surgical training time, and 22 (62.9%) reported a perceived decrease in training quality. Simulated skills training was only available to eight (22.9%) participants. Twenty-four (68.6%) experienced burnout and/or depression symptoms during the pandemic. Twenty-seven (77.1%) reported that postgraduate research was unaffected by the pandemic. CONCLUSION: During the COVID-19 pandemic, surgical trainees at this institution reported a decrease in the quality of surgical training and skills acquisition and a negative impact on their mental wellbeing.

Surgical Catastrophic Health Expenditure and Risk Factors for Out-of-Pocket Expenditure at a South African Public Sector Hospital.

BACKGROUND: Worldwide, 3.7 billion people risk financial catastrophe if they require surgery, mostly affecting the poorest populations. Surgical care associated with catastrophic health expenditure (CHE) has not been well-described in the South African context. The objectives of this study were to determine: (1) the proportion of surgical patients at a South African hospital who experienced CHE and impoverishing health expenditure (IHE); and (2) the risk factors for out-of-pocket (OOP) payments. METHODS: A cross-sectional prospective questionnaire was administered to participants admitted for a surgical procedure at New Somerset Hospital, Cape Town. CHE was defined in three ways: (1) 40% or more of capacity-to-pay, (2) 25% of annual household expenditure, or (3) 10% of annual household expenditure. IHE was described as the number of participants who experienced new or worsening impoverishment after surgery. RESULTS: Two hundred and seventy-four participants were interviewed, and 263 were included in the analysis (4% attrition rate). Two (0.8%) participants experienced CHE. 98.5% of participants spent less than 10% of their annual household expenditure and 43 participants (16.7%) experienced IHE. Risk factors for OOP expenditure were cancer diagnosis (p = 0.0386), an elective procedure (p = 0.0001), and having a limited health insurance plan (p = 0.0492). DISCUSSION: Most participants undergoing a surgical procedure did not experience CHE. Participants were relatively protected from financial catastrophe owing to subsidized user fees and the provision of transport. However, 17% of patients experienced IHE, suggesting even small payments resulted in impoverishment. Ensuring low financial vulnerabilities around surgical care is an important consideration for national surgical planning in South Africa.

Establishing a South African national framework for COVID-19 surgical prioritisation.

BACKGROUND: Since the start of the COVID-19 pandemic, surgical operations have been drastically reduced in South Africa (SA). Guidelines on surgical prioritisation during COVID-19 have been published, but are specific to high-income countries. There is a pressing need for context-specific guidelines and a validated tool for prioritising surgical cases during the COVID-19 pandemic. In March 2020, the South African National Surgical Obstetric Anaesthesia Plan Task Team was asked by the National Department of Health to establish a national framework for COVID-19 surgical prioritisation. OBJECTIVES: To develop a national framework for COVID-19 surgical prioritisation, including a set of recommendations and a risk calculatorfor operative care. METHODS: The surgical prioritisation framework was developed in three stages: (i) a literature review of international, national and local recommendations on COVID-19 and surgical care was conducted; (ii) a set of recommendations was drawn up based on the available literature and through consensus of the COVID-19 Task Team; and (iii) a COVID-19 surgical risk calculator was developed and evaluated. RESULTS: A total of 30 documents were identified from which recommendations around prioritisation of surgical care were used to draw up six recommendations for preoperative COVID-19 screening and testing as well as the use of appropriate personal protective equipment. Ninety-nine perioperative practitioners from eight SA provinces evaluated the COVID-19 surgical risk calculator, which had high acceptability and a high level of concordance (81%) with current clinical practice. CONCLUSIONS: This national framework on COVID-19 surgical prioritisation can help hospital teams make ethical, equitable and personalised decisions whether to proceed with or delay surgical operations during this unprecedented epidemic.

Estimating the surgical backlog from the COVID-19 lockdown in South Africa: A retrospective analysis of six government hospitals.

BACKGROUND: The COVID-19 pandemic reached South Africa (SA) in March 2020. A national lockdown began on 27 March 2020, and health facilities reduced non-essential activity, including many surgical services. PRIMARY OBJECTIVE: to estimate the COVID-19 surgical backlog in Western Cape Province, SA, by comparing 2019 and 2020 general surgery operative volume and proportion at six district and regional hospitals. SECONDARY OBJECTIVE: to compare the operative volume of appendicectomy, laparoscopic cholecystectomy, cancer and trauma between the 2 years. METHODS: This was a retrospective study of general surgery operations from six SA government hospitals in the Western Cape. Data were obtained from electronic operative databases or operative theatre logbooks from 1 April to 31 July 2019 and 1 April to 31 July 2020. RESULTS: Total general surgery operations decreased by 44% between 2019 (n=3 247) and 2020 (n=1 810) (p<0.001). Elective operations decreased by 74% (n=1 379 v. n=362; p<0.001), and one common elective procedure, laparoscopic cholecystectomy, decreased by 68% (p<0.001). Emergency operations decreased by 22% (n=1 868 v. n=1 448; p<0.001) and trauma operations by 42% (n=325 v. n=190; p<0.001). However, non-trauma emergency operations such as appendicectomy and cancer did not decrease. The surgical backlog for elective operations after 4 months from these six hospitals is 1 017 cases, which will take between 4 and 14 months to address if each hospital can do one additional operation per weekday. CONCLUSIONS: The COVID-19 pandemic has created large backlogs of elective operations that will need to be addressed urgently. Clear and structured guidelines need to be developed in order to streamline the reintroduction of full surgical healthcare services as SA slowly recovers from this unprecedented pandemic.

District hospital surgical capacity in Western Cape Province, South Africa: A cross-sectional survey.

BACKGROUND: The role of the district hospital (DH) in surgical care has been undervalued. However, decentralised surgical services at DHs have been identified as a key component of universal health coverage. Surgical capacity at DHs in Western Cape (WC) Province, South Africa, has not been described. OBJECTIVES: To describe DH surgical capacity in WC and identify barriers to scaling up surgical capacity at these facilities. METHODS: This was a cross-sectional survey of 33 DHs using the World Health Organization surgical situational analysis tool administered to hospital staff from June to December 2019. The survey addressed the following domains: general services and financing; service delivery and surgical volume; surgical workforce; hospital and operating theatre (OT) infrastructure, equipment and medication; and barriers to scaling up surgical care. RESULTS: Seven of 33 DHs (21%) did not have a functional OT. Of the 28 World Bank DH procedures, small WC DHs performed up to 22 (79%) and medium/large DHs up to 26 (93%). Only medium/large DHs performed all three bellwether procedures. Five DHs (15%) had a full-time surgeon, anaesthetist or obstetrician (SAO). Of DHs without any SAO specialists, 14 (50%) had family physicians (FPs). These DHs performed more operative procedures than those without FPs (p=0.005). Lack of finances dedicated for surgical care and lack of surgical providers were the most reported barriers to providing and expanding surgical services. CONCLUSIONS: WC DH surgical capacity varied by hospital size. However, FPs could play an essential role in surgery at DHs with appropriate training, oversight and support from SAO specialists. Strategies to scale up surgical capacity include dedicated financial and human resources.

Changes in surgical practice in 85 South African hospitals during COVID-19 hard lockdown.

BACKGROUND: In preparation for the COVID-19 pandemic, South Africa (SA) began a national lockdown on 27 March 2020, and many hospitals implemented measures to prepare for a potential COVID-19 surge. OBJECTIVES: To report changes in SA hospital surgical practices in response to COVID-19 preparedness. METHODS: In this cross-sectional study, surgeons working in SA hospitals were recruited through surgical professional associations via an online survey. The main outcome measures were changes in hospital practice around surgical decision-making, operating theatres, surgical services and surgical trainees, and the potential long-term effect of these changes. RESULTS: A total of 133 surgeons from 85 hospitals representing public and private hospitals nationwide responded. In 59 hospitals (69.4%), surgeons were involved in the decision to de-escalate surgical care. Access was cancelled or reduced for non-cancer elective (n=84; 99.0%), cancer (n=24; 28.1%) and emergency operations (n=46; 54.1%), and 26 hospitals (30.6%) repurposed at least one operating room as a ventilated critical care bed. Routine postoperative visits were cancelled in 33 hospitals (36.5%) and conducted by telephone or video in 15 (16.6%), 74 hospitals (87.1%) cancelled or reduced new outpatient visits, 64 (75.3%) reallocated some surgical inpatient beds to COVID-19 cases, and 29 (34.1%) deployed some surgical staff (including trainees) to other hospital services such as COVID-19 testing, medical/COVID-19 wards, the emergency department and the intensive care unit. CONCLUSIONS: Hospital surgical de-escalation in response to COVID-19 has greatly reduced access to surgical care in SA, which could result in a backlog of surgical needs and an excess of morbidity and mortality.

Changes in surgical practice in 85 South African hospitals during COVID-19 hard lockdown

Background. In preparation for the COVID-19 pandemic, South Africa (SA) began a national lockdown on 27 March 2020, and many hospitals implemented measures to prepare for a potential COVID-19 surge.Objectives. To report changes in SA hospital surgical practices in response to COVID-19 preparedness.Methods. In this cross-sectional study, surgeons working in SA hospitals were recruited through surgical professional associations via an online survey. The main outcome measures were changes in hospital practice around surgical decision-making, operating theatres, surgical services and surgical trainees, and the potential long-term effect of these changes.Results. A total of 133 surgeons from 85 hospitals representing public and private hospitals nationwide responded. In 59 hospitals (69.4%), surgeons were involved in the decision to de-escalate surgical care. Access was cancelled or reduced for non-cancer elective (n=84; 99.0%), cancer (n=24; 28.1%) and emergency operations (n=46; 54.1%), and 26 hospitals (30.6%) repurposed at least one operating room as a ventilated critical care bed. Routine postoperative visits were cancelled in 33 hospitals (36.5%) and conducted by telephone or video in 15 (16.6%), 74 hospitals (87.1%) cancelled or reduced new outpatient visits, 64 (75.3%) reallocated some surgical inpatient beds to COVID-19 cases, and 29 (34.1%) deployed some surgical staff (including trainees) to other hospital services such as COVID-19 testing, medical/COVID-19 wards, the emergency department and the intensive care unit.Conclusions. Hospital surgical de-escalation in response to COVID-19 has greatly reduced access to surgical care in SA, which could result in a backlog of surgical needs and an excess of morbidity and mortality

Molecular biomarkers for prognosis of gastrointestinal stromal tumor.

Gastrointestinal stromal tumor (GIST) is the most common mesenchymal tumor of the gastrointestinal tract. However, the development of molecular markers, especially circulating biomarkers, remains largely undone for the prognosis of GIST. We discussed the clinical-pathological characteristics of GIST and identified potential biomarkers for guidance of therapy and prognosis of GIST. Around 90% of GISTs contain mutations in KIT or PDGFRA and the remaining 10% of GISTs are wild-type. Recent studies have indicated that various DNAs and miRNAs could serve as potential biomarkers for prognosis of GIST, including KIT, PDGFRA, other DNAs (such as BRAF, SDH, SETD2 and ROR2), and microRNAs (miRNAs). The pressing need and challenges in the development of circulating prognostic biomarkers for GIST are also discussed. Although challenges remain, DNAs and miRNAs are promising circulating biomarkers for surveillance and prognosis of GIST. Advances in clarification of aberrant molecular alterations may open new avenues for exploration of reliable and robust biomarkers to improve the management of GIST.

The value of neck drain in esophageal surgery: a randomized trial.

The use of surgical drains in certain clean elective operations remains controversial. To evaluate the role of closed-suction drain for an esophageal anastomosis in the neck, we conducted a randomized, controlled study in 40 patients with esophageal carcinoma who underwent esophagectomy with an esophageal anastomosis in the neck, half of whom had a neck drain inserted at the end of operation. The median (range) duration of drainage was 46 hours (36 to 88 hours). The median (range) amount of drainage was 63 ml (15 to 210 ml). There was no incidence of haematoma or seroma formation in both the drained and non-drained groups. Anastomotic leakage did not occur in any patient. The benefits of closed suction neck drain could not be demonstrated. Routine use of neck drain for esophageal anastomosis in the neck is not necessary.

Circulating cell-free DNAs and miRNAs as promising non-invasive biomarkers for early detection of gastric cancer.

Early diagnosis of gastric cancer is critical to decrease the mortality of this globally fatal disease. Currently, endoscopic biopsy is the gold standard for diagnosis of gastric cancer. However, invasiveness and high expense limit its application. Hence, non-invasive and cost-effective biomarkers for early detection and subsequent management are crucial steps to monitor gastric cancer. Recent studies suggest that circulating cell-free nucleic acids, including circulating tumor DNAs and microRNAs (miRNAs), are promising for various applications. Development of such blood-based biomarkers is expected to facilitate detection, predict prognosis, monitor chemotherapeutic response and manage recurrence of human cancers. In this review, the characteristics of circulating cell-free DNAs and miRNAs will be elucidated, including their origin and dysregulation. Mutations and hypermethylation of circulating DNAs, abnormal alternations of expression of circulating miRNAs will be revealed as aberrant changes indicating gastric cancer. The roles of circulating DNAs and miRNAs for early detection of gastric cancer will be focused on, as well as the challenges of developing circulating nucleic acids as biomarkers.

Modeling the effects of cyclodextrin on intracellular membrane vesicles from Cos-7 cells prepared by sonication and carbonate treatment.

Cholesterol has important functions in the organization of membrane structure and this may be mediated via the formation of cholesterol-rich, liquid-ordered membrane microdomains often referred to as lipid rafts. Methyl-beta-cyclodextrin (cyclodextrin) is commonly used in cell biology studies to extract cholesterol and therefore disrupt lipid rafts. However, in this study we reassessed this experimental strategy and investigated the effects of cyclodextrin on the physical properties of sonicated and carbonate-treated intracellular membrane vesicles isolated from Cos-7 fibroblasts. We treated these membranes, which mainly originate from the trans-Golgi network and endosomes, with cyclodextrin and measured the effects on their equilibrium buoyant density, protein content, represented by the palmitoylated protein phosphatidylinositol 4-kinase type IIα, and cholesterol. Despite the reduction in mass stemming from cholesterol removal, the vesicles became denser, indicating a possible large volumetric decrease, and this was confirmed by measurements of hydrodynamic vesicle size. Subsequent mathematical analyses demonstrated that only half of this change in membrane size was attributable to cholesterol loss. Hence, the non-selective desorption properties of cyclodextrin are also involved in membrane size and density changes. These findings may have implications for preceding studies that interpreted cyclodextrin-induced changes to membrane biochemistry in the context of lipid raft disruption without taking into account our finding that cyclodextrin treatment also reduces membrane size.

Hospital Authority audit of the outcome of endoscopic resection of superficial upper gastro-intestinal lesions in Hong Kong.

OBJECTIVES: To review the short-term outcome of endoscopic resection of superficial upper gastro-intestinal lesions in Hong Kong. DESIGN: Historical cohort study. SETTING: All Hospital Authority hospitals in Hong Kong. PATIENTS: This was a multicentre retrospective study of all patients who underwent endoscopic resection of superficial upper gastro-intestinal lesions between January 2010 and June 2013 in all government-funded hospitals in Hong Kong. MAIN OUTCOME MEASURES: Indication of the procedures, peri-procedural and procedural parameters, oncological outcomes, morbidity, and mortality. RESULTS: During the study period, 187 lesions in 168 patients were resected. Endoscopic mucosal resection was performed in 34 (18.2%) lesions and endoscopic submucosal dissection in 153 (81.8%) lesions. The mean size of the lesions was 2.6 (standard deviation, 1.8) cm. The 30-day morbidity rate was 14.4%, and perforations and severe bleeding occurred in 4.3% and 3.2% of the patients, respectively. Among patients who had dysplasia or carcinoma, R0 resection was achieved in 78% and the piecemeal resection rate was 11.8%. Lateral margin involvement was 14% and vertical margin involvement was 8%. Local recurrence occurred in 9% of patients and 15% had residual disease. The 2-year overall survival rate and disease-specific survival rate was 90.6% and 100%, respectively. CONCLUSION: Endoscopic mucosal resection and endoscopic submucosal dissection were introduced in low-to-moderate-volume hospitals with acceptable morbidity rates. The short-term survival was excellent. However, other oncological outcomes were higher than those observed in high-volume centres and more secondary procedures were required.

Intranasal dexmedetomidine in combination with patient-controlled sedation during upper gastrointestinal endoscopy: a randomised trial.

BACKGROUND: Sedation using intranasal dexmedetomidine is a convenient and well-tolerated technique. This study evaluated the sedative efficacy of intranasal dexmedetomidine in combination with patient-controlled sedation (PCS) for upper gastrointestinal endoscopy. METHODS: In this double-blind, randomised, controlled trial, 50 patients received either intranasal dexmedetomidine 1.5 µg/kg (dexmedetomidine group) or intranasal saline (placebo group) 1 h before the procedure. PCS with propofol and alfentanil was provided for rescue sedation. Additional sedative consumption, perioperative sedation scores using Observer's Assessment of Alertness/Sedation (OAA/S) scale, recovery, vital signs, adverse events and patient satisfaction were assessed. RESULTS: Total consumption of PCS propofol and alfentanil was significantly less in the dexmedetomidine than placebo group with a mean difference of -13.8 mg propofol (95% confidence interval -27.3 to -0.3) and -34.5 µg alfentanil (95% confidence interval -68.2 to -0.7) at the completion of the procedure (P = 0.044). Weighted areas under the curve (AUCw ) of OAA/S scores were significantly lower in the dexmedetomidine group before, during and after procedures (P < 0.001, P = 0.024 and P = 0.041 respectively). AUCw of heart rate and systolic blood pressure were also significantly lower during the procedure (P = 0.007 and P = 0.022 respectively) with dexmedetomidine. There was no difference in recovery, side effects or satisfaction. CONCLUSION: Intranasal dexmedetomidine with PCS propofol and alfentanil confers deeper perioperative clinical sedation with significantly less use of additional sedatives during upper gastrointestinal endoscopy.

Nevirapine- and efavirenz-associated hepatotoxicity under programmatic conditions in Kenya and Mozambique.

To describe the frequency, risk factors, and clinical signs and symptoms associated with hepatotoxicity (HT) in patients on nevirapine- or efavirenz-based antiretroviral therapy (ART), we conducted a retrospective cohort analysis of patients attending the ART clinic in Kibera, Kenya, from April 2003 to December 2006 and in Mavalane, Mozambique, from December 2002 to March 2007. Data were collected on 5832 HIV-positive individuals who had initiated nevirapine- or efavirenz-based ART. Median baseline CD4+ count was 125 cells/µL (interquartile range [IQR] 55-196). Over a median follow-up time of 426 (IQR 147-693) days, 124 (2.4%) patients developed HT. Forty-one (54.7%) of 75 patients with grade 3 HT compared with 21 (80.8%) of 26 with grade 4 had associated clinical signs or symptoms (P = 0.018). Four (5.7%) of 124 patients with HT died in the first six months compared with 271 (5.3%) of 5159 patients who did not develop HT (P = 0.315). The proportion of patients developing HT was low and HT was not associated with increased mortality. Clinical signs and symptoms identified 50% of grade 3 HT and most cases of grade 4 HT. This suggests that in settings where alanine aminotransferase measurement is not feasible, nevirapine- and efavirenz-based ART may be given safely without laboratory monitoring.

Detergent-free isolation and characterization of cholesterol-rich membrane domains from trans-Golgi network vesicles.

Cholesterol is an abundant lipid of the trans-Golgi network (TGN) and of certain endosomal membranes where cholesterol-rich microdomains are important in the organization and compartmentalization of vesicular trafficking. Here we describe the development of a rapid method to isolate a cholesterol-rich endomembrane fraction. We show that widely used subcellular fractionation techniques incompletely separate cholesterol-rich membranes, such as the TGN, from organelles, such as late endosomes and lysosomes. To address this issue, we devised a new subcellular fractionation scheme involving two rounds of velocity centrifugation, membrane sonication, and discontinuous sucrose density gradient centrifugation. This strategy resulted in the isolation of a cholesterol and GM1 glycosphingolipid-enriched membrane fraction that was completely cleared of plasma membrane, endoplasmic reticulum, and mitochondria. This buoyant fraction was enriched for the TGN and recycling endosome proteins Rab11 and syntaxin-6, and it was well resolved from cis-Golgi and early and late endosomal membranes. We demonstrate that this technique can give useful insights into the compartmentation of phosphoinositide synthesis, and it facilitates the isolation of cholesterol-rich membranes from a population of TGN-trafficking vesicles.

Relationship between phosphatidylinositol 4-phosphate synthesis, membrane organization, and lateral diffusion of PI4KIIalpha at the trans-Golgi network.

Type II phosphatidylinositol 4-kinase IIalpha (PI4KIIalpha) is the dominant phosphatidylinositol kinase activity measured in mammalian cells and has important functions in intracellular vesicular trafficking. Recently PI4KIIalpha has been shown to have important roles in neuronal survival and tumorigenesis. This study focuses on the relationship between membrane cholesterol levels, phosphatidylinositol 4-phosphate (PI4P) synthesis, and PI4KIIalpha mobility. Enzyme kinetic measurements, sterol substitution studies, and membrane fragmentation analyses all revealed that cholesterol regulates PI4KIIalpha activity indirectly through effects on membrane structure. In particular, we found that cholesterol levels determined the distribution of PI4KIIalpha to biophysically distinct membrane domains. Imaging studies on cells expressing enhanced green fluorescent protein (eGFP)-tagged PI4KIIalpha demonstrated that cholesterol depletion resulted in morphological changes to the juxtanuclear membrane pool of the enzyme. Lateral membrane diffusion of eGFP-PI4KIIalpha was assessed by fluorescence recovery after photobleaching (FRAP) experiments, which revealed the existence of both mobile and immobile pools of the enzyme. Sterol depletion decreased the size of the mobile pool of PI4KIIalpha. Further measurements revealed that the reduction in the mobile fraction of PI4KIIalpha correlated with a loss of trans-Golgi network (TGN) membrane connectivity. We conclude that cholesterol modulates PI4P synthesis through effects on membrane organization and enzyme diffusion.

Differential effects of the phosphatidylinositol 4-kinases, PI4KIIα and PI4KIIIß, on Akt activation and apoptosis.

In this study, we investigated the role of PI4P synthesis by the phosphatidylinositol 4-kinases, PI4KIIα and PI4KIIIß, in epidermal growth factor (EGF)-stimulated phosphoinositide signaling and cell survival. In COS-7 cells, knockdown of either isozyme by RNA interference reduced basal levels of PI4P and PI(4,5)P(2), without affecting receptor activation. Only knockdown of PI4KIIα inhibited EGF-stimulated Akt phosphorylation, indicating that decreased PI(4,5)P(2) synthesis observed by loss of either isoform could not account for this PI4KIIα-specific effect. Phospholipase Cγ activation was also differentially affected by knockdown of either PI4K isozyme. Overexpression of kinase-inactive PI4KIIα, which induces defective endosomal trafficking without reducing PI(4,5)P(2) levels, also reduced Akt activation. Furthermore, PI4KIIα knockdown profoundly inhibited cell proliferation and induced apoptosis as evidenced by the cleavage of caspase-3 and its substrate poly(ADP-ribose) polymerase. However, in MDA-MB-231 breast cancer cells, apoptosis was observed subsequent to knockdown of either PI4KIIα or PI4KIIIß and this correlated with enhanced proapoptotic Akt phosphorylation. The differential effects of phosphatidylinositol 4-kinase knockdown in the two cell lines lead to the conclusion that phosphoinositide turnover is inhibited through PI4P substrate depletion, whereas impaired antiapoptotic Akt signaling is an indirect consequence of dysfunctional endosomal trafficking.

Enhanced nitric oxide and cyclic GMP formation plays a role in the anti-platelet activity of simvastatin.

BACKGROUND AND PURPOSE: It has been found that 3-hydroxy-3-methyl-glutaryl coenzyme A (HMG-CoA) reductase inhibitors (statins) exert various vascular protective effects, beyond their cholesterol-lowering property, including inhibition of platelet-dependent thrombus formation. The objective of the present study was to determine whether the nitric oxide (NO)/cyclic GMP-mediated processes in platelets contribute to the anti-aggregatory activity of simvastatin. EXPERIMENTAL APPROACH: After rabbit platelets were incubated with simvastatin for 5 min, aggregation was induced and the platelet aggregation, nitric oxide synthase activity, guanylyl cyclase activity, NO and cyclic GMP formation were measured appropriately. KEY RESULTS: Treatment with simvastatin concentration-dependently inhibited platelet aggregation induced by collagen or arachidonic acid with an IC(50) range of 52-158 microM. We also demonstrated that simvastatin (20-80 microM) concentration-dependently further enhanced collagen-induced NO and cyclic GMP formation through increasing NOS activity (from 2.64+/-0.12 to 3.52+/-0.21-5.10+/-0.14 micromol min(-1) mg protein(-1)) and guanylyl cyclase activity (from 142.9+/-7.2 to 163.5+/-17.5-283.8+/-19.5 pmol min(-1) mg protein(-1)) in the platelets. On the contrary, inhibition of platelet aggregation by simvastatin was markedly attenuated (by about 50%) by addition of a nitric oxide synthase inhibitor, a NO scavenger or a NO-sensitive guanylyl cyclase inhibitor. The anti-aggregatory effects of simvastatin were significantly increased by addition of a selective inhibitor of cyclic GMP phosphodiesterase. CONCLUSIONS AND IMPLICATIONS: Our findings indicate that enhancement of a NO/cyclic GMP-mediated process plays an important role in the anti-aggregatory activity of simvastatin.