Minimal information for studies of extracellular vesicles (MISEV2023): From basic to advanced approaches

Extracellular vesicles (EVs), through their complex cargo, can reflect the state of their cell of origin and change the functions and phenotypes of other cells. These features indicate strong biomarker and therapeutic potential and have generated broad interest, as evidenced by the steady year-on-year increase in the numbers of scientific publications about EVs. Important advances have been made in EV metrology and in understanding and applying EV biology. However, hurdles remain to realising the potential of EVs in domains ranging from basic biology to clinical applications due to challenges in EV nomenclature, separation from non-vesicular extracellular particles, characterisation and functional studies. To address the challenges and opportunities in this rapidly evolving field, the International Society for Extracellular Vesicles (ISEV) updates its 'Minimal Information for Studies of Extracellular Vesicles', which was first published in 2014 and then in 2018 as MISEV2014 and MISEV2018, respectively. The goal of the current document, MISEV2023, is to provide researchers with an updated snapshot of available approaches and their advantages and limitations for production, separation and characterisation of EVs from multiple sources, including cell culture, body fluids and solid tissues. In addition to presenting the latest state of the art in basic principles of EV research, this document also covers advanced techniques and approaches that are currently expanding the boundaries of the field. MISEV2023 also includes new sections on EV release and uptake and a brief discussion of in vivo approaches to study EVs. Compiling feedback from ISEV expert task forces and more than 1000 researchers, this document conveys the current state of EV research to facilitate robust scientific discoveries and move the field forward even more rapidly

Postprandial reactive hypoglycaemia: varying presentation patterns on extended glucose tolerance tests and possible therapeutic approaches.

Reactive hypoglycemia is a state characterised by sympathetic or neuroglycopenic symptoms associated with hypoglycaemia in the postprandial state resulting in considerable distress to the patient. It is our practice to carry out either extended glucose tolerance tests (eGTTs) or mixed meal tests in these patients. We describe two patients who experienced hypoglycaemic symptoms early and late during eGTT. The patient who experienced symptoms early, in contrast to the patient who presented with late symptoms, did not possess any characteristics of the metabolic syndrome. Based on clinical symptoms, glucose, insulin, and free fatty acid (FFA) levels, we speculate on possible mechanisms that may have accounted for each of their presentation patterns. We then discuss low glycaemic index diet which will be the mainstay of management

Fibrates and estimated glomerular filtration rate: observations from an outpatient clinic setting and clinical implications.

BACKGROUND Previous studies have demonstrated that fibrates have an effect on creatinine concentrations. The pattern of change with fibrates in estimated glomerular filtration rate (eGFR), widely used in clinical practice, has not been previously described. METHODS Data was retrospectively collected from 132 consecutive case notes of patients started on fibrates in a lipid clinic between 2002 and 2008. Pre- and post-fibrate creatinine concentrations were measured and eGFR measurements were obtained. RESULTS Of the 79 patients with both pre and post-treatment eGFR values 8 ml/min/1.73 m(2), 25% demonstrated a reduction >16 ml/min/1.73 m(2), and 10% demonstrated a reduction >21 ml/min/1.73 m(2). CONCLUSIONS The authors demonstrate a significant effect of fibrates on eGFR in clinical practice. Awareness of the pattern of eGFR change is important for decisions regarding the continued use of fibrate therapy and/or commonly co-prescribed diabetic drugs and renal specialist referrals

Testosterone replacement therapy with long-acting testosterone undecanoate improves sexual function and quality-of-life parameters vs. placebo in a population of men with type 2 diabetes.

INTRODUCTION Sexual dysfunction, particularly erectile dysfunction (ED), is common in men with type 2 diabetes, occurring in up to 75% of cases. The prevalence of hypogonadism is also high in men with diabetes and low testosterone is associated with both sexual dysfunction and a reduced response to oral therapy for ED. AIM This study aimed to determine the effect of testosterone replacement with long-acting Testosterone Undecanoate (TU) on sexual function, mood and quality of life vs. placebo over a treatment period of 30 weeks followed by 52 weeks of open-label medication. The study was conducted in a primary care population of men with type 2 diabetes attending their primary care physician for routine visits. METHODS The male diabetic populations of seven general practices were screened at routine diabetes visits to detect symptomatic men with total testosterone levels of 12 nmol/L or less or with free testosterones of 250 pmol/L or less. Two hundred eleven men were screened. A double-blind placebo-controlled study was conducted in 199 men with type 2 diabetes and hypogonadism treated for 30 weeks with either 1,000 mg of TU or matching placebo followed by 52-week open-label follow on. MAIN OUTCOME MEASURES The primary outcome measure, International Index of Erectile Function (IIEF), was used to evaluate sexual dysfunction, and the Ageing Male Symptom (AMS), Hospital Anxiety and Depression Scale, and Global Efficacy Question were used as secondary outcome measures to assess mood and self-reported quality of life. RESULTS Testosterone replacement therapy with long-acting TU improved all domains of sexual function at 30 weeks (erectile function [EF], P = 0.005; intercourse satisfaction, P = 0.015; sexual desire, P = 0.001; overall satisfaction, P = 0.05; and orgasm, P = 0.04), with benefit as early as 6 weeks. Improvements in AMS score were significant in men without depression (P = 0.02) and the presence of depression at baseline was associated with marked reduction in response to both sexual function and psychological scores. All responses in sexual function continued to improve significantly up to 18 months with an improvement in EF score of 4.31 from baseline. In a small cohort of 35 men taking phosphodiesterase type 5 inhibitors, there was no change during the double-blind phase but a nine-point improvement in EF domain during 52-week open-label treatment. After 30 weeks, 46% vs. 17% of patients on active therapy vs. placebo felt that the treatment had improved their health, reaching 70% after open-label therapy. Less obese and older patients responded better to testosterone therapy. There were no significant adverse events. CONCLUSION TU significantly improved all domains of the IIEF and patient reported quality of life at 30 weeks and more significantly after 52-week open-label extension. Improvement was most marked in less obese patient and those without coexisting depression. In men with type 2 diabetes, trials of therapy may need to be given for much longer than 3-6 months suggested in current guidelines

Testosterone replacement therapy improves metabolic parameters in hypogonadal men with type 2 diabetes but not in men with coexisting depression: the BLAST study.

INTRODUCTION The association between testosterone deficiency and insulin resistance in men with type 2 diabetes is well established and current endocrine society guidelines recommend the measurement of testosterone levels in all men with type 2 diabetes or erectile dysfunction. AIM We report the first double-blind, placebo-controlled study conducted exclusively in a male type 2 diabetes population to assess metabolic changes with long-acting testosterone undecanoate (TU). METHODS The type 2 diabetes registers of seven general practices identified 211 patients for a 30-week double-blind, placebo-controlled study of long-acting TU 1,000 mg followed by 52 weeks of open-label use. Because of the established impact of age, obesity, and depression on sexual function, these variables were also assessed for influence on metabolic parameters. MAIN OUTCOME MEASURE Changes in glycated hemoglobin (HbA1c) and the level of testosterone at which response are achieved. RESULTS Treatment with TU produced a statistically significant reduction in HbA1c at 6 and 18 weeks and after a further 52 weeks of open-label medication most marked in poorly controlled patients with baseline HbA1c greater than 7.5 where the reduction was 0.41% within 6 weeks, and a further 0.46% after 52 weeks of open-label use. There was significant reduction in waist circumference, weight, and body mass index in men without depression, and improvements were related to achieving adequate serum levels of testosterone. There were no significant safety issues. CONCLUSIONS Testosterone replacement therapy significantly improved HbA1c, total cholesterol, and waist circumference in men with type 2 diabetes. Improvements were less marked in men with depression at baseline, and therapeutic responses were related to achieving adequate serum testosterone levels. Current advice on 3- to 6-month trials of therapy may be insufficient to achieve maximal response. Patients reported significant improvements in general health

Effect of fibrate treatment on liver function tests in patients with the metabolic syndrome.

BACKGROUND Fibrates are used especially in patients with hypertriglyceridaemia, a feature of the metabolic syndrome. Elevated LFTs are often observed in these patients perhaps related to fatty infiltration. AIM We wished to study changes seen in LFTs (GGT, ALT and ALP) following fibrate therapy and then determine associated factors. METHODS This was a retrospective observational study in which data was collected from case notes of patients started on fibrates (n = 118, 2002-2008) in the lipid clinic at Good Hope Hospital and pre/post-fibrate lipid and LFT values were obtained. All biochemistry was performed on the Roche P-Unit using supplied reagents. Statistical analyses included t tests and regression analyses (factorised when quartiles were compared). RESULTS Of the study population 106 patients were on fenofibrate; the remaining on bezafibrate. Significant lowering of GGT (p  57 IU/l, ALT > 34 IU/l and ALP > 94 IU/l) significantly different to the lowest quartile. The above associations remained significant even when the regression analyses were corrected for changes in lipid values (which did not show an association). CONCLUSIONS Fibrate treatment led to improvements in LFT, the greatest benefit seen in patients with higher baseline LFT values. It appears that baseline and changes in lipid values post fibrate treatment were not associated with change in LFT

Significant increase in high-density lipoprotein cholesterol with fibrates is associated with low pretreatment high-density lipoprotein cholesterol: findings from an outpatient clinic setting.

BACKGROUND Change in high-density lipoprotein cholesterol (HDL-C) observed in large randomized controlled trials using fibrates has varied. Inconsistent cardiovascular outcomes have also been the common theme of these trials. Subgroup analysis of even the negative trials, however, reveals significant reduction in cardiovascular disease in patients with low HDL-C and high triglycerides. We wished to study HDL-C change following fibrate therapy in our lipid clinic and determine the factors associated with HDL-C change. METHODS Data were collected from case notes of patients started on fibrates (n=248) between 2002 and 2008 in the lipid clinics at Heart of England NHS Foundation Trust. Regression analyses were carried out to determine factors associated with changes in HDL-C. RESULTS Linear regression analysis revealed that HDL-C change was associated with pretreatment HDL-C (P<0.001), diabetes (P=0.004) and treatment duration (P=0.036). Multiple regression analysis with all of the factors in the model suggested that they were independent. Patients with a baseline HDL-C <1.0 mmol/L showed a greater HDL-C increase when compared to patients with a baseline HDL-C ≥1.0 mmol/L; HDL-C <1.0 mmol/L (increase of 0.15 mmol/L, linear regression: c=0.14, 95% confidence interval 0.05-0.30, P<0.001) and HDL-C ≥1.0 (increase of 0.002 mmol/L, linear regression: reference category). A similar relationship between change in HDL-C and baseline HDL-C was observed within groups stratified by patient characteristics (apart from those on concurrent statin therapy and females). CONCLUSIONS Our results may explain the discrepancies observed in some randomized controlled trials whereby subgroup analysis of patients with the metabolic syndrome appeared to show benefit whereas this was absent in the total cohort. Thus, future interventional studies using fibrates should perhaps focus on patients with low HDL-C levels

The response to testosterone undecanoate in men with type 2 diabetes is dependent on achieving threshold serum levels (the BLAST study).

BACKGROUND The association between testosterone deficiency and insulin resistance in men with type 2 diabetes is well established. Current Endocrine Society and European Association of Urology guidelines recommend the measurement of testosterone levels in all men with type 2 diabetes and in men suffering from erectile dysfunction. It is recognised that a range of physical symptoms appear as the testosterone level falls but few studies have addressed the threshold at which symptoms improve with physiological replacement. We report the first double-blind placebo-controlled study conducted exclusively in a male type 2 diabetes population to assess the metabolic changes with testosterone replacement. METHODS The type 2 diabetes registers of seven general practices were screened to establish the prevalence of low testosterone and the associations with diabetes control. Of 550 eligible patients approached, 488 men (mean age 62.6) consented to take part in screening with a morning testosterone level, assessed between 8 and 11 am. This identified 211 patients for a double-blind placebo-controlled study of long acting testosterone undecanoate (TU) 1000 mg lasting 30 weeks followed by 52 weeks of open label use. The population was divided into a SEVERE group with either total testosterone (TT) of 8 nmol/l or less or free testosterone (FT) 180 pmol/l or less or a MILD group with TT 8.1-12 nmol/l or FT 181-250 pmol/l. RESULTS Men in the SEVERE group increased mean through TT from 7.73 nmol/l at baseline to 9.93 at 30 weeks and the MILD group from 10.47 to 11.94. The SEVERE group showed marked improvement in sexual function, but no significant improvement in metabolic parameters. The MILD group showed no improvement in sexual function, but significant improvement in weight, body mass index, waist circumference and Hospital Anxiety and Depression Scale. Improvement was seen in all parameters during 52 weeks open label treatment where trough TT levels approached 15 nmol/l. Baseline prostate-specific antigen (PSA) was lower in the SEVERE group and increased with TU for 30 weeks and then stabilised. There was no increase in PSA with treatment in the MILD group. CONCLUSIONS Testosterone undecanoate significantly improves sexual parameters and Ageing Male Symptom Score, but not metabolic factors at 30 weeks in men with SEVERE testosterone deficiency syndrome (TDS). In men with MILD TDS, significant improvements in metabolic but not sexual parameters were seen, suggesting that there are threshold levels for response to testosterone replacement therapy and that trials of therapy need to achieve sustained therapeutic levels to be effective. PSA showed minor rises, but only for 30 weeks in the SEVERE group