Time to thrombolysis and factors contributing to delays in patients presenting with ST-elevation myocardial infarction at Chris Hani Baragwanath Academic Hospital, Johannesburg, South Africa.

BACKGROUND: Acute coronary syndrome is a public health burden both worldwide and in South Africa (SA). Guidelines recommend thrombolysis within 1 hour of symptom onset and 30 minutes of hospital arrival for patients with ST-elevation myocardial infarction (STEMI) in order to prevent morbidity and mortality. There is a paucity of data pertaining to the time between onset of chest pain and thrombolysis in STEMI patients in SA. OBJECTIVES: To elucidate the time to thrombolytic therapy, establish the reasons for treatment delays, and calculate the loss of benefit of thrombolysis associated with delays in treatment of patients presenting with STEMI at Chris Hani Baragwanath Academic Hospital (CHBAH), Johannesburg, SA. METHOD: A prospective observational study of 100 consecutive patients with STEMI was conducted at CHBAH (2021 - 2022). RESULTS: The mean (standard deviation) age was 55.6 (11.6) years, with a male predominance (78%). Thrombolytic therapy was administered to 51 patients, with a median (interquartile range (IQR)) time to thrombolysis of 360 (258 - 768) minutes; 10 of the patients who received a thrombolytic (19.6%) did so within 30 minutes of arrival at the hospital. The median (IQR) time from symptom onset to calling for help was 60 (30 - 240) minutes, the median time from arrival of help to hospital arrival was 114 (48 - 468) minutes, and the median in-hospital delay to thrombolysis after arrival was 105 (45 - 240) minutes. Numerous reasons that led to delay in treatment were identified, but the most frequent was prehospital delays related to patient factors. Late presentation resulted in 26/49 patients (53.1%) not receiving thrombolytic therapy. Five patients died and 43 suffered from heart failure. Thirty per 1 000 participants could have been saved had they received thrombolytic therapy within 1 hour from the onset of chest pain. CONCLUSION: Prehospital and hospital-related factors played a significant role in delays to thrombolysis that led to increased morbidity and mortality of patients with STEMI.

SA Heart consensus statement on closure of patent foramen ovale 2021.

A patent foramen ovale (PFO) is associated with numerous clinical conditions. The most severe of these is cryptogenic stroke. This consensus statement aims to provide a clinical guideline on which patients should be offered PFO closure.

Association of TYRP1 with hypoxia and its correlation with patient outcome in uveal melanoma

Purpose Molecular mechanisms of uveal melanoma development in association with high pigmentation are unclear. Tyrosinase Related Protein (TYRP1) is not only one of the important melanogenesis marker that contributes to melanin synthesis, but can also prevents the melanocyte death. The induction of melanogenesis leads to induction of HIF-1α which can affect the behavior of melanoma cells and its surrounding environment. The aim of our study was to determine the expression of TYRP1 and HIF-1α at the protein and RNA level and determine its prognostic significance. Methods In the present study, the expression of TYRP1 and HIF-1α was investigated on 61 formalin-fixed paraffin-embedded choroidal melanoma samples by immunohistochemistry. Fresh 50 samples were validated by real-time PCR. Results were correlated with clinicopathological parameters and Kaplan–Meier was performed to determine the prognostic significance. Results High immunoexpression of TYRP1 and HIF-1α was present in 61 and 54% of patients, respectively. Both TYRP1 and HIF-1α correlated well with high pigmentation and BAP1 (BRCA1 Associated Protein-1) loss (p < 0.05) at IHC level as well as transcriptional level. There was reduced metastatic free survival in patients with necrosis and this was statistically significant (p = 0.010). Conclusion Our findings indicate that TYRP1 can be used as a potential biomarker in the development of targeted therapy in UM. Further studies on melanogenesis markers associated with TYRP1 could provide us a better understanding in this field (AU)

Association of TYRP1 with hypoxia and its correlation with patient outcome in uveal melanoma.

PURPOSE: Molecular mechanisms of uveal melanoma development in association with high pigmentation are unclear. Tyrosinase Related Protein (TYRP1) is not only one of the important melanogenesis marker that contributes to melanin synthesis, but can also prevents the melanocyte death. The induction of melanogenesis leads to induction of HIF-1α which can affect the behavior of melanoma cells and its surrounding environment. The aim of our study was to determine the expression of TYRP1 and HIF-1α at the protein and RNA level and determine its prognostic significance. METHODS: In the present study, the expression of TYRP1 and HIF-1α was investigated on 61 formalin-fixed paraffin-embedded choroidal melanoma samples by immunohistochemistry. Fresh 50 samples were validated by real-time PCR. Results were correlated with clinicopathological parameters and Kaplan-Meier was performed to determine the prognostic significance. RESULTS: High immunoexpression of TYRP1 and HIF-1α was present in 61 and 54% of patients, respectively. Both TYRP1 and HIF-1α correlated well with high pigmentation and BAP1 (BRCA1 Associated Protein-1) loss (p < 0.05) at IHC level as well as transcriptional level. There was reduced metastatic free survival in patients with necrosis and this was statistically significant (p = 0.010). CONCLUSION: Our findings indicate that TYRP1 can be used as a potential biomarker in the development of targeted therapy in UM. Further studies on melanogenesis markers associated with TYRP1 could provide us a better understanding in this field.

Constitutive expression of c-REL in uveal melanoma patients: correlation with clinicopathological parameters and patient outcome.

PURPOSE: Uveal melanoma (UM) is the most common intraocular cancer with a high mortality rate that requires new research in the field of prevention and treatment. c-REL is a member of the nuclear factor κB (NF-κB) transcription factor family and an emerging regulator of tumorigenesis. Therefore, the objective of the study is to evaluate the constitutive expression of c-REL in uveal melanoma patients and its prognostic significance. METHODS: Detection of c-REL expression was carried out by immunohistochemistry in all 75 patients, and qRT-PCR performed on 58 fresh cases of uveal melanoma along with IL-6 status. Immunoblot was performed to validate immunohistochemistry results. Expression of c-REL protein correlated with clinicopathological parameters and overall survival of patients. RESULTS: Immunohistochemistry results revealed nuclear expression of the c-REL protein (56%) in our cases. Out of 75 cases, 31 cases showed nuclear expression, and 11 cases had cytoplasmic expression. qRT-PCR showed upregulation of the REL gene in 56.89% cases at the transcriptional level. There was a statistically significant difference in the overall survival of patients with c-REL nuclear immunopositivity (p = 0.0048). On multivariate analysis, scleral invasion and c-REL nuclear expression found to be an independent prognostic factor (p < 0.05) CONCLUSIONS: To the best of our knowledge, this was the first study reporting the expression of the c-REL protein in uveal melanoma. Strong nuclear immunoexpression of c-Rel suggests NFκB pathway activation which might be involved in the progression of the disease. Differential expression of c-REL protein may be used as an attractive target for the development of anticancer strategies.

Sensitivity of MG-ADL for generalized weakness in myasthenia gravis.

BACKGROUND AND PURPOSE: Myasthenia gravis activities of daily living (MG-ADL) is a commonly used questionnaire in MG trials. To investigate whether MG-ADL is equally sensitive to oculobulbar and generalized weakness, its correlation with the oculobulbar and generalized domain of the quantitative myasthenia gravis (QMG) score was analyzed (QMGob and QMGgen, respectively). To test whether the sensitivity of MG-ADL for generalized weakness could be improved, the additional value of ACTIVLIM on top of MG-ADL in the prediction QMGgen in was investigated. METHODS: MG-ADL, QMG and ACTIVLIM, an ADL questionnaire focusing on generalized weakness, were analyzed in a prospective cohort of 112 MG patients. A generalized linear model was used to calculate the correlation of MG-ADL with QMGob and QMGgen and to assess the additional value of ACTIVLIM on top of MG-ADL for its correlation with QMGgen. RESULTS: MG-ADL had a higher correlation with QMGob than with QMGgen (B = 0.68, P < 0.001, and B = 0.38, P < 0.001, respectively). A similar trend was found for changes in the scores (B = 0.68, P = 0.132, and B = 0.39, P = 0.492, respectively). ACTIVLIM had a significant additional value on top of MG-ADL in the prediction of QMGgen, both cross-sectionally (B = -0.61, P < 0.001) and for changes within individual patients (B = -0.93, P = 0.041). CONCLUSION: MG-ADL has a lower sensitivity for generalized weakness than for oculobulbar weakness. Adding questions on generalized weakness would improve the sensitivity of the MG-ADL for generalized weakness.

Does NEMO/IKKgamma protein have a role in determining prognostic significance in uveal melanoma?

Purpose: Uveal melanoma, although a rare form of cancer, is the most common primary malignancy of the eye in adults. Nuclear factor-kapaB (NF-kapaB) is a transcription factor that transactivates genes involved in the regulation of cell growth, apoptosis, angiogenesis, and metastasis, but the molecular mechanisms that negatively regulate NF-kapaB activation are not fully understood. NF-kapaB can also be activated by DNA damage pathway through NEMO protein. Therefore, the objective of this study is to elucidate the role of NEMO/IKKgamma protein in uveal melanoma patients. Methods: Seventy-five formalin-fixed paraffin-embedded prospective tissues of uveal melanoma were included in the present study. These cases were reviewed and investigated for the expression of NEMO/IKKgamma protein by immunohistochemistry and validated by western blotting along with the qRT-PCR for mRNA expression. Expression levels were correlated with the clinicopathological parameters and patients’ outcome. Results: Immunohistochemistry showed cytoplasmic expression of NEMO/IKKgamma expression in only 22 out of 75 (29.33%) cases. This result was confirmed by western blotting, and correlated well with the immunohistochemical expression of NEMO/IKKgamma protein (48 kDa). In addition, downregulation of this gene was found in 87.93% of the cases when compared with the normal tissues. On statistical analysis, loss of NEMO/IKKgamma protein was correlated with neovascularization, high mitotic count, and presence of vascular loop (p < 0.05). There was less overall survival rate with low expression of NEMO/IKKgamma protein in patients with uveal melanoma. Conclusion: This was the first study suggesting the relevant role of NEMO/IKKgamma protein, and highlights the prognostic significance with outcome in uveal melanoma patients. This protein might be used as a screening biomarker in these patients after large-scale validation and translational studies

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Striking increase in the incidence of infective endocarditis associated with recreational drug abuse in urban South Africa.

BACKGROUND: We have recently noted a dramatic rise in the number of patients with infective endocarditis (IE) related to intravenous (IV) nyaope (a mixture of heroin, cocaine and antiretroviral drugs) presenting to Chris Hani Baragwanath Academic Hospital in Johannesburg, South Africa. OBJECTIVES: To document the clinical and echocardiographic characteristics of these patients. METHODS: Clinical and echocardiographic characteristics of all patients (N=68) with IE due to IV nyaope use were retrospectively extracted from hospital records (December 2014 - February 2017). RESULTS: The mean (standard deviation) age of the patients was 25.8 (4.5) years (97.1% were male). Withdrawal symptoms were noted in 25.1% of cases, fever in 58.8%, dyspnoea in 86.7% and right ventricular failure in 42.6%. Most patients were HIV-positive (76.1%), with CD4+ cell counts of <200 cells/µL in 8.8% of the total, 58.1% had hepatitis C infection, and only three were on antiretrovirals. Septic pulmonary emboli were noted in 61.8%. Blood cultures revealed Staphylococcus aureus in 61.2%, Enterococcus faecalis in 8.8% and Pseudomonas aeruginosa in 1 patient; 29.2% had sterile cultures and 8.8% polymicrobial infection. Severe right ventricular systolic dysfunction (RVS' Doppler velocity <10 cm/s) and pulmonary hypertension were noted in 19.1% and 62.2% of patients, respectively. Pericardial effusion was noted in 37.8%. The most commonly involved valve was the tricuspid (60.1%), followed by the mitral (17.2%), aortic (2.9%) and pulmonary (1 patient) valves. Combined valve lesions were noted in 19.1% of patients. Ten patients (14.7%) died. The main predictor of in-hospital mortality was S. aureus infection (odds ratio 5.0; p=0.042). CONCLUSIONS: We have documented the common clinical and echocardiographic characteristics of patients with IE secondary to IV nyaope use. IE due to IV drug use is responsible for considerable morbidity and mortality in a predominantly young male population.

Does NEMO/IKKγ protein have a role in determining prognostic significance in uveal melanoma?

PURPOSE: Uveal melanoma, although a rare form of cancer, is the most common primary malignancy of the eye in adults. Nuclear factor-κB (NF-κB) is a transcription factor that transactivates genes involved in the regulation of cell growth, apoptosis, angiogenesis, and metastasis, but the molecular mechanisms that negatively regulate NF-κB activation are not fully understood. NF-κB can also be activated by DNA damage pathway through NEMO protein. Therefore, the objective of this study is to elucidate the role of NEMO/IKKγ protein in uveal melanoma patients. METHODS: Seventy-five formalin-fixed paraffin-embedded prospective tissues of uveal melanoma were included in the present study. These cases were reviewed and investigated for the expression of NEMO/IKKγ protein by immunohistochemistry and validated by western blotting along with the qRT-PCR for mRNA expression. Expression levels were correlated with the clinicopathological parameters and patients' outcome. RESULTS: Immunohistochemistry showed cytoplasmic expression of NEMO/IKKγ expression in only 22 out of 75 (29.33%) cases. This result was confirmed by western blotting, and correlated well with the immunohistochemical expression of NEMO/IKKγ protein (48 kDa). In addition, downregulation of this gene was found in 87.93% of the cases when compared with the normal tissues. On statistical analysis, loss of NEMO/IKKγ protein was correlated with neovascularization, high mitotic count, and presence of vascular loop (p < 0.05). There was less overall survival rate with low expression of NEMO/IKKγ protein in patients with uveal melanoma. CONCLUSION: This was the first study suggesting the relevant role of NEMO/IKKγ protein, and highlights the prognostic significance with outcome in uveal melanoma patients. This protein might be used as a screening biomarker in these patients after large-scale validation and translational studies.

PEGylated and targeted extracellular vesicles display enhanced cell specificity and circulation time.

Extracellular vesicles (EVs) are increasingly being recognized as candidate drug delivery systems due to their ability to functionally transfer biological cargo between cells. However, the therapeutic applicability of EVs may be limited due to a lack of cell-targeting specificity and rapid clearance of exogenous EVs from the circulation. In order to improve EV characteristics for drug delivery to tumor cells, we have developed a novel method for decorating EVs with targeting ligands conjugated to polyethylene glycol (PEG). Nanobodies specific for the epidermal growth factor receptor (EGFR) were conjugated to phospholipid (DMPE)-PEG derivatives to prepare nanobody-PEG-micelles. When micelles were mixed with EVs derived from Neuro2A cells or platelets, a temperature-dependent transfer of nanobody-PEG-lipids to the EV membranes was observed, indicative of a 'post-insertion' mechanism. This process did not affect EV morphology, size distribution, or protein composition. After introduction of PEG-conjugated control nanobodies to EVs, cellular binding was compromised due to the shielding properties of PEG. However, specific binding to EGFR-overexpressing tumor cells was dramatically increased when EGFR-specific nanobodies were employed. Moreover, whereas unmodified EVs were rapidly cleared from the circulation within 10min after intravenous injection in mice, EVs modified with nanobody-PEG-lipids were still detectable in plasma for longer than 60min post-injection. In conclusion, we propose post-insertion as a novel technique to confer targeting capacity to isolated EVs, circumventing the requirement to modify EV-secreting cells. Importantly, insertion of ligand-conjugated PEG-derivatized phospholipids in EV membranes equips EVs with improved cell specificity and prolonged circulation times, potentially increasing EV accumulation in targeted tissues and improving cargo delivery.

Impression cytology diagnosis of ulcerative eyelid malignancy.

OBJECTIVE: The utility of impression cytology in ocular diseases has predominantly been restricted to the diagnosis of dry eye, limbal stem cell deficiency and conjunctival neoplasias. Its role in malignant eyelid lesions remains largely unexplored. Although scrape cytology is more popular for cutaneous lesions, impression cytology, being non-traumatic, has an advantage in small and delicate areas such as the eyelid. The present study has been designed to evaluate its role in the diagnosis and management of malignant eyelid lesions. METHODS: Thirty-two histopathologically proven malignant eyelid lesions diagnosed over a 2-year period, including 13 basal cell carcinomas, 11 sebaceous carcinomas, four squamous cell carcinomas, two malignant melanomas and two poorly differentiated carcinomas, formed the study group. RESULTS: The results of impression cytology were compared with those of histopathology in the study group and with an age- and sex-matched group of benign cases as controls. The sensitivity of impression cytology was 84% (27/32) for the diagnosis of malignancy and 28% (9/32) for categorization of the type of malignancy. CONCLUSIONS: Impression cytology is a simple, useful, non-invasive technique for the detection of malignant ulcerative eyelid lesions. It is especially useful as a follow-up technique for the detection of recurrences.

Toward routine detection of extracellular vesicles in clinical samples.

The majority, if not all, of human cell types secrete extracellular vesicles (EVs) into their environment, at least partly as a means of intercellular communication. These secreted vesicles can be detected in most bodily fluids including blood, urine, and saliva. The number of secreted vesicles and their composition is altered in various pathological conditions, raising opportunities to exploit EVs as diagnostic and/or prognostic biomarkers. For this to become a reality, it is important to reach consensus regarding the standardization of protocols for sample collection, EV isolation, handling, and storage for valid comparison and interpretation of measurements. Depending on the information required, there are several detection options including EV number and size distribution, molecular surface markers, procoagulation activity, and RNA content. For these purposes, different techniques are currently utilized or under development. This review discusses the techniques that have the potential to become standard EV detection methods in a clinical diagnostic setting. In addition to the accuracy of the detection technique, other factors such as high-throughput, cost-effectiveness, time consumption, and required operator skill are important to consider. A combination of increasing fundamental knowledge, technological progress, standardization of sample collection, and processing protocols is required for EVs to become reliable predictors of altered physiology or development of disease suitable for routine clinical diagnostics. Cancer and (cardio)vascular disorders are examples of pathologies where EV detection may be applied in the near future for diagnosis and/or prognosis.

Tricuspid valve endocarditis associated with intravenous nyoape use: a report of 3 cases.

We report three cases of tricuspid valve infective endocarditis associated with intravenous nyoape use. Nyoape is a variable drug combination of an antiretroviral (efavirenz or ritonavir), heroin, metamphetamines and cannabis. Its use is becoming increasingly common among poor communities in South Africa. All our patients were young HIV-positive men from disadvantaged backgrounds. They all presented with tricuspid regurgitation and septic pulmonary emboli. They were treated with prolonged intravenous antibiotic courses, and one required referral for surgery.

Targeted delivery of small interfering RNA to angiogenic endothelial cells with liposome-polycation-DNA particles.

Angiogenesis is an attractive target for cancer therapy, due to its central position in tumor growth and development. Vascular Endothelial Growth Factor (VEGF) and its receptors (VEGFRs) play a key role in the angiogenic process. A promising strategy for targeting VEGF-mediated angiogenesis is RNA interference (RNAi) using short interfering RNA (siRNA). However, for efficacious RNAi a well-designed siRNA delivery system is crucial. Liposome-Polycation-DNA (LPD) particles form a promising system for siRNA delivery to tumors. In order to target angiogenic endothelial cells, LPD particles may be modified with a targeting ligand, such as a cyclic Arg-Gly-Asp (RGD) peptide that specifically binds to integrins expressed on tumor-associated endothelial cells. In the current study, RGD-targeted PEGylated LPD particles containing VEGFR-2 siRNA were prepared and optimized with respect to their size and charge by varying protamine content, carrier DNA content for stronger complexation, and PEGylation density. The size of the optimized particles was around 200 nm and the ζ-potential was approximately +20 mV. The uptake and silencing efficacy of the RGD-targeted PEGylated LPD particles were evaluated in H5V cells (murine endothelial cells) and Human Umbilical Vein Endothelial cells (HUVECs). When compared to non-targeted LPD particles, enhanced uptake and silencing of VEGFR-2 expression was observed for RGD-targeted PEGylated LPD particles. In conclusion, the RGD-targeted PEGylated LPD particles containing VEGFR-2 siRNA presented here may be a promising approach for targeting VEGF-mediated angiogenesis in cancer therapy.

Examining the role of Rac1 in tumor angiogenesis and growth: a clinically relevant RNAi-mediated approach.

Angiogenesis, the sprouting of new blood vessels from the pre-existing vasculature, is a well established target in anti-cancer therapy. It is thought that the Rho GTPase Rac1 is required during vascular endothelial growth factor (VEGF)-mediated angiogenesis. In the present study, we have used a clinically relevant RNA interference approach to silence Rac1 expression. Human umbilical vein endothelial cells were transiently transfected with non-specific control siRNA (siNS) or Rac1 siRNA (siRac1) using electroporation or Lipofectamine 2000. Functional assays with transfected endothelial cells were performed to determine the effect of Rac1 knockdown on angiogenesis in vitro. Silencing of Rac1 inhibited VEGF-mediated tube formation, cell migration, invasion and proliferation. In addition, treatment with Rac1 siRNA inhibited angiogenesis in an in vivo Matrigel plug assay. Intratumoral injections of siRac1 almost completely inhibited the growth of grafted Neuro2a tumors and reduced tumor angiogenesis. Together, these data indicate that Rac1 is an important regulator of VEGF-mediated angiogenesis. Knockdown of Rac1 may represent an attractive approach to inhibit tumor angiogenesis and growth.