Current dissociation methods for solid tissues in scRNA-seq studies do not guarantee intact single-cell isolation, especially for sensitive and complex human endocrine tissues. Most studies rely on enzymatic dissociation of fresh samples or nuclei isolation from frozen samples. Dissociating whole intact cells from fresh-frozen samples, commonly collected by biobanks, remains a challenge. Here, we utilized the acetic-methanol dissociation approach (ACME) to capture transcriptional profiles of individual cells from fresh-frozen tissue samples. This method combines acetic acid-based dissociation and methanol-based fixation. In our study, we optimized this approach for human endocrine tissue samples for the first time. We incorporated a high-salt washing buffer instead of the standard PBS to stabilize RNA and prevent RNases reactivation during rehydration. We have designated this optimized protocol as ACME HS (ACetic acid-MEthanol High Salt). This technique aims to preserve cell morphology and RNA integrity, minimizing transcriptome changes and providing a more accurate representation of mature mRNA. We compared the ability of enzymatic, ACME HS, and nuclei isolation methods to preserve major cell types, gene expression, and standard quality parameters across 41 tissue samples. Our results demonstrated that ACME HS effectively dissociates and fixes cells, preserving cell morphology and high RNA integrity. This makes ACME HS a valuable alternative for scRNA-seq protocols involving challenging tissues where obtaining a live cell suspension is difficult or disruptive.

IntroductionEndocrine system disorders are a serious public health burden and can be caused by deleterious genetic variants in single genes or by the combined effects of multiple variants along with environmental and lifestyle factors.MethodsThe EndoGene database presents the results of next-generation sequencing assays used to genetically profile 5,926 patients who were diagnosed with 450 endocrine and concomitant diseases and were examined and treated at the National Medical Research Center for Endocrinology between November 2017 and January 2024. Among them, 494, 1,785, 692, and 1,941 patients were profiled using four internally developed genetic panels including 220, 250, 376, and 382 genes, respectively, selected based on a literature analysis and clinical recommendations, and 1,245 patients were profiled by whole exome sequencing covering 31,969 genes.Results2,711 genetic variants were reported as clinically relevant by medical geneticists and are presented here along with genomic, technical, and clinical annotations.DiscussionThis publicly accessible database will be useful to those interested in genetics, epidemiology, population statistics, and a better understanding of the molecular basis of endocrine disorders.

BACKGROUND: Diabetes mellitus (DM) is a dangerous challenge to society and high-priority goals for health systems worldwide due to the severity of diabetic complications leading to disability and premature mortality in this category of patients. AIM: To carry out an analysis of epidemiological, clinical and therapeutic characteristics of DM control in the Russian Federation (RF) according to the strategy goals of the World Health Organization (WHO) by 2030 (diagnosis of DM; control of glycemia, blood pressure in DM patients; assignment of statins to DM patients; ensuring the availability of insulin and means for self-monitoring of the level of glycemia) based on the << Database of clinical and epidemiological monitoring of DM in the territory of the Russian Federation (RF)>>. MATERIALS AND METHODS. The research performed on the basis of the de-personalized "Database of clinical and epidemiological monitoring of DM in the territory of the RF", certificate of state registration database N degrees 2020622447 (http://diaregistry.ru, register of DM), which included 87 regions of the RF. Data are presented as cross-sectional study in 01.07.2024. RESULTS: According to the Database of clinical and epidemiological monitoring of DM in the territory of the RF, the target HbA(1c)<7% is achieved in 29% of patients with type 1 DM (T1DM) and 42% of patients with T2DM. Effective blood pressure control in people with diagnosed DM is achieved in 84% of patients with T1DM and 60% of patients with T2DM. The proportion of patients receiving statins aged 40 and over is 14.1% for T1DM, 15.9% for T2DM. All patients with T1DM (100%) and T2DM on insulin therapy are provided with insulin under the program of state guarantee of drug provision, of them insulin analogues in T1DM - 84,8%, in T2DM - 60,9%, the rest - recombinant human insulins. Self-monitoring devices (glucometers and test strips for them) are provided to 100% of patients with DM. Since 2023, patients with T1DM under 18 years old are provided by continuous glucose monitoring systems. CONCLUSION: Common diabetes monitoring system based on "Database of clinical and epidemiological monitoring of DM in the territory of the RF" at the national level allows to assess the effectiveness of programs to achieve the WHO targets. The data analysis identifies priority areas for the most effective and achievable interventions to strengthen and monitor diabetes control. Providing timely, reliable and sustainable national information on risk factors, epidemiology and clinical data of DM, access to medical drugs and modern technologies of control requires coordinated work by health professionals.

Chronic kidney disease (CKD) is a supranosological concept, defined as a disorder of the structure and/or function of the kidneys for 3 months or more that leads to health consequences. CKD develops in 20–40% of patients with diabetes mellitus (DM). Dynamic analysis of epidemiological indicators of CKD in patients with diabetes mellitus points to the prevalence increase of this pathology in the Russian Federation in 2010–2022. Improved quality of diagnostics of earlier stages complications, a decrease in the risk of cardiovascular events (the leading cause of death) and end-stage renal failure at a later age and with a longer duration of diabetes are noted as well. The development of tools for predicting the risk of the pathology progression and its complications has made it possible to improve patient monitoring and routing. Current international and domestic recommendations emphasize a comprehensive approach to the treatment of CKD, including lifestyle modification, evidence-based pharmacological treatments aimed at preserving organ function, and improving cardiovascular prognosis. An optimal model of care for patients with diabetes and CKD, aimed at multidisciplinary care and adapted according to the severity of CKD, will lead to the best outcomes for the individual patient and society.

Omics technologies accumulated an enormous amount of data that advanced knowledge about the molecular pathogenesis of type 1 diabetes mellitus and identified a number of fundamental problems focused on the transition to personalized diabetology in the future. Among them, the most significant are the following: (1) clinical and genetic heterogeneity of type 1 diabetes mellitus; (2) the prognostic significance of DNA markers beyond the HLA genes; (3) assessment of the contribution of a large number of DNA markers to the polygenic risk of disease progress; (4) the existence of ethnic population differences in the distribution of frequencies of risk alleles and genotypes; (5) the infancy of epigenetic research into type 1 diabetes mellitus. Disclosure of these issues is one of the priorities of fundamental diabetology and practical healthcare. The purpose of this review is the systemization of the results of modern molecular genetic, transcriptomic, and epigenetic investigations of type 1 diabetes mellitus in general, as well as its individual forms. The paper summarizes data on the role of risk HLA haplotypes and a number of other candidate genes and loci, identified through genome-wide association studies, in the development of this disease and in alterations in T cell signaling. In addition, this review assesses the contribution of differential DNA methylation and the role of microRNAs in the formation of the molecular pathogenesis of type 1 diabetes mellitus, as well as discusses the most currently central trends in the context of early diagnosis of type 1 diabetes mellitus.

Dear Colleagues!We are glad to present the 11th Edition of the Standards of Specialized Diabetes Care. These evidence-based guidelines were designed to standardize and facilitate diabetes care in all regions of the Russian Federation.The Standards are updated on the regular basis to incorporate new data and relevant recommendations from national and international clinical societies, including World Health Organization Guidelines (WHO), International Diabetes Federation (IDF), European Association for the Study of Diabetes (EASD), American Diabetes Association (ADA), American Association of Clinical Endocrinologists (AACE), International Society for Pediatric and Adolescent Diabetes (ISPAD) and Russian Association of Endocrinologists (RAE). Current edition of the “Standards” also integrates results of completed randomized clinical trials, as well as findings from the national studies of diabetes mellitus (DM), conducted in close partnership with a number of Russian hospitals.Latest data indicates that prevalence of DM in the world increased during the last decade more than two-fold, reaching some 537 million patients by the end of 2021. According to the current estimation by the International Diabetes Federation, 643 million patients will be suffering from DM by 2030 and 783 million by 2045.Like many other countries, Russian Federation experiences a sharp rise in the prevalence of DM. According to Russian Federal Diabetes Register, there are at least 4 962 762 patients with DM in this country on 01.01.2023 (3,42% of population) with 92,3% (4 581 990) – Type 2 DM, 5,6% (277 092) – Type 1 DM and 2,1% (103 680) – other types of DM, including 8 758 women with gestational DM. However, these results underestimate real quantity of patients, because they consider only registered cases. Results of Russian epidemiological study (NATION) confirmed that 54% of patients with Type 2 DM are undiagnosed. So real number of patients with DM in Russia is 11-12 million patients (about 7% of population). This is a great long-term problem, because a lot of patients are not diagnosed, so they don’t receive any treatment and have high risk of vascular complications.Severe consequences of the global pandemic of DM include its vascular complications: nephropathy, retinopathy, coronary, cerebral and peripheral vascular disease. These conditions are responsible for the majority of cases of diabetes-related disability and death.In accordance with the Federal Law № 489-FZ of 25.12.2018 medical care should be provided on the basis of the clinical recommendations. Therefore, clinical recommendations posted in the rubricator of the Ministry of Health of the Russian Federation (https://cr.minzdrav.gov.ru) being the priority.This edition of the Algorithms for Specialized Care for Patients with Diabetes Mellitus contains the main information from the clinical guidelines for the management of diabetes mellitus in adults and children (“Type 1 Diabetes in Children (2022)”, “Type 1 Diabetes in Adults (2022)”, “Type 2 diabetes mellitus in children (2021)”, “Type 2 diabetes mellitus in adults (2022)”), as well as the updated data.New information has been added about indications for using of antihyperglycemic drugs and their instructional restrictions; algorithm of treatment of type 2 diabetes mellitus has been updated; clarifications have been made on the range of glycemic control in pregnant women; information on the treatment of diabetic complications and the management of pediatric patients has been added. In 2023, Russian diabetology suffered an unbearable loss - Alexander Yuryevich Mayorov, who was the executive editor of the previous issues and an active participant of the work on all the previous editions of “Algorithms” from the moment they were founded, passed away. This publication is dedicated to the memory of Alexander Yuryevich, who was an example of the service to patients and medicine, a friend, teacher, mentor, generously sharing his knowledge and the sample of the execution of the medical duty to all of us. On behalf of the Working Group

Background: Bariatric surgery is known to induce weight loss and diabetes remission in patients with type 2 diabetes (T2D), but the exact mechanism of glycemic normalization needs to be defined. Methods: The study included patients with BMI ≥ 35 kg/m2, obesity history ≥ 10 years, and planned bariatric surgery. At baseline and 3 and 6 months after surgery, all patients underwent anthropometric measurements, body composition and blood tests (including insulin, glucagon, and incretins during oral glucose tolerance test (OGTT)), and hyperinsulinemic euglycemic clamp tests. Diabetes remission was defined if the person reached HbA1c < 6.5% after surgery and glucose-lowering therapy withdrawal. Results: The study included 86 patients, divided into groups with no diabetes (control group, n = 44) and T2D (n = 42). Most patients with T2D reached normoglycemia at 6 months. BMI and insulin resistance (according to M-index) decreased in T2D group comparably to people without diabetes. At 6 months, people with T2D at baseline had less insulin and GLP-1 secretion and higher glucagon level during OGTT when compared to the control group. Conclusions: We conclude that weight and insulin resistance reduction is sufficient for T2D remission. The absence of insulin, glucagon, and incretin restoration is not crucial for the glucose metabolism in the short-term, but it may explain the relapse of T2D years after bariatric surgery.

We recommend acromegaly to be ruled in all patients with characteristic changes in appearance (A3). In all patients without characteristic changes in appearance, we recommend to rule out acromegaly, if several clinical signs suspicious for acromegaly are present (B3). We recommend acromegaly to be ruled out in all patients with pituitary adenoma (B3). We recommend to rule out acromegaly in all patients with prolactin-secreting adenomas (C4). For laboratory confirmation of acromegaly, we recommend to measure serum insulin-like growth factor-1 (IGF-1, somatomedin C) level (A3). In patients without obvious clinical signs of acromegaly and/or with a moderate increase in IGF-1 levels (IGF-1 index less than 1.3), we recommend to assess the response of somatotropic hormone (GH) to hyperglycemia (GH during an oral glucose tolerance test) (B3). In all patients with biochemically confirmed acromegaly, magnetic resonance imaging of the pituitary gland with intravenous contrast is recommended to assess pituitary adenoma size, location and growth pattern (A3). We recommend to measure blood glucose and glycated hemoglobin in all patients with confirmed acromegaly (B3). We recommend to measure serum prolactin levels in all patients with acromegaly (B3).In all patients with acromegaly and pituitary adenoma, transnasal transsphenoidal adenomectomy is recommended as first-line treatment if the patient’s consent is given and there are no contraindications (A1). For all patients with acromegaly for whom neurosurgical treatment is indicated, surgical intervention is recommended in medical centers, specializing in pituitary diseases, by neurosurgeons who perform at least 50 such operations per year (A3). We recommend that morphological and immunohistochemical examination of the removed pituitary adenoma is performed in all patients with acromegaly (A3). We do not recommend routine medical therapy for acromegaly before surgery as a mean to reduce tumor size (B2).For patients with acromegaly in whom neurosurgical treatment is not indicated or ineffective or while patient awaiting for the effect of radiation therapy, we recommend medical therapy (A1). Long-acting first-generation somatostatin analogues are recommended as first-line drug therapy for acromegaly. In case of partial/complete resistance to or intolerance to somatostatin analogs, pegvisomant is recommended as a second-line medical therapy for acromegaly (A1). We recommend starting pegvisomant from a single dose of 40–80 mg per day subcutaneously, then continue injections at a starting dose of 10 or 15 mg per day with IGF-1 monitoring after 4–6 weeks and dose adjustment if necessary (B2). In patients with acromegaly and a moderate increase in IGF-1 levels, we recommend to initiate cabergoline as medical monotherapy or, if somatostatin analogs are ineffective, in combination with them (B2).In patients with acromegaly, if the disease remains active after surgical treatment, if drug treatment is unavailable, intolerable or ineffective, and repeated neurosurgical intervention is inappropriate, it is we recommend radiation therapy (A1). A multidisciplinary approach is recommended in the management of acromegaly complications (C3).

Aim. To study the immediate efficacy, tolerability and predictors of response to prolonged-acting somatostatin analogues (SSAs) in patients with ectopic ACTH syndrome (EAS). Materials and methods. A multicenter, observational study with a retrospective analysis. The effectiveness of treatment was evaluated every 12–24 weeks by the activity of hypercortisolism, clinical symptoms of the disease, control of tumor growth. Patients were conditionally divided into “responders” and “non-responders” at time points of 6, 12 and 24 months. Radiological efficacy was performed according to the RECIST 1.1. Statistical data processing was carried out by using IBM SPSS Statistics 23. Results. The study included 46 patients (26 women, 20 men). The median follow-up period was 71 months [32; 122]. Localized tumor process (T1-3N0M0) – in 19 (41.3%) patients, locally widespread (T1-3N1M0, T4N0-1M0) – in 15 (32.6%), generalized (T1-4N0-1M1) – in 12 (26.1%) cases. Surgical treatment was performed in 33 (71.7%) patients. SSAs were the first-line therapy for all 46 patients. The average duration of SSAs use was 34 months (Me 27.5 [13.8; 41]). Dose escalation was required in 39.4% cases, the period before the first dose escalation was on average 9 months. 13 (48.1%) patients achieved drug-induced remission of the disease. Normalization of cortisol in 24hUFC or its decrease ≥50% from the baseline level by the 6th month of treatment was achieved in 46.4%, by the 12th month – in 61.9%, “eluding” from the effect of therapy was observed in 3 patients. Objective response was evaluated in 30 patients. There was no complete response to the treatment. In 4 (13.3%) cases – a partial response, in 15 (50%) – disease stabilization, in 11 (36.7%) – disease progression. The best therapeutic efficacy of SSAs is associated with bronchial NET (HR 0.078, 95% CI 0.010–0.633; p=0.017), and resistance to treatment is associated with negative expression of SSTR5 (HR 6.532, 95% CI 1.019–41.878; p=0.048). Conclusion. SSAs are the drugs of choice for the first-line treatment of patients with EAS, allowing for long-term effective control of hypercortisolism and tumor progression in more than 60% of patients. Significant factors determining the response to SSAs after 6 months of treatment are expression status of SSTR5 and NET localization.

AIM: The purpose of this study was to analyze the characteristics of management, diagnosis and treatment of patients with vitamin D deficiency/ by endocrinologists in the regions of Russian Federation using a sociological survey.MATERIALS AND METHODS: A cross-sectional sociological uncontrolled study was carried out. To implement this work, we used an online questionnaire “Questioning doctors on vitamin D” specially developed on the basis of the Google forms platform. The study was conducted in January 2021.All the data obtained were entered into an electronic database in MS Excel. Statistical processing was performed using the IBM SPSS Statistics 25 software.RESULTS: The survey involved 707 physicians from 86 regions of the Russian Federation. Our study revealed that not all doctors strictly adhere to clinical recommendations in patient management. Issues identified include unjustifiably widespread ordering of 25(OH)D blood tests (58.5%), lack of consideration for individual patient characteristics and clinical situations in dose selection (99.2%) and prescription of active metabolite or analogs of vitamin D to correct low 25(OH)D levels in blood (14.7%). Furthermore, recommendations for improving clinical guidelines focused on the lack of illustrative material (21.1%), expanding patient information sections (20.7%), and insufficient coverage of issues arising in clinical practice (14.6%). Our study also highlighted limited capabilities of laboratory diagnostics for assessing vitamin D status in patients in Russia. The survey revealed that only 9.2% of respondents have the ability to measure 1,25(OH)2D concentrations, and only 1.4% can order tests for 24,25(OH)2D. About 8.3% of participants did not specify which tests for assessing vitamin D status are available for prescription. Technical enhancement of laboratories and the inclusion of all recommended laboratory study requirements in the compulsory health insurance system could address this.Regarding vitamin D toxicity, 25% of surveyed doctors encountered it. Main causes included self-administration of elevated doses of cholecalciferol by patients (62%) or prescribed by physicians (40%), the use of active metabolites or analogs of vitamin D (21%), incorrect dosing of cholecalciferol preparations as indicated by the manufacturer (18%), and defects in CYP24A1 (13%). Rare causes included granulomatous and lymphoproliferative diseases (11.5%).CONCLUSION: The current clinical guidelines of the Russian Association of Endocrinologists for «Vitamin D Deficiency in Adults» are generally effective and widely used by clinicians. However, they do not entirely meet the needs of specialists and require refinement in terms of improving structure and clarity of presentation, expanding illustrative material, and justifying the frequency of laboratory diagnostics. Cases of vitamin D toxicity observed in clinical practice were predominantly associated with incorrect administration of vitamin D supplements. The identified high frequency of vitamin D toxicity in real clinical practice necessitates additional attention to this issue during the update of clinical recommendations.

Current scRNA-seq studies of solid tissues mostly rely on enzymatic dissociation of fresh samples or the fallback on nuclei isolation from frozen or partially fixed samples. However, due to the complex tissue organization or cell fragility, it could be challenging to apply these approaches to the sensitive endocrine tissues. That is, dissociating intact cells from such problematic fresh-frozen samples routinely collected by biobanks remains challenging.In this study, we adapted the acetic-methanol dissociation method – ACME High Salt (ACME HS) to effectively isolate intact single cells from fresh-frozen endocrine tumor samples, including adrenal gland neoplasms, thyroid carcinomas, and pituitary neuroendocrine tumors. We compared the ability of enzymatic, ACME HS, and nuclear isolation methods to preserve the integrity of major cell types and gene expression across 41 tissue samples of different origins. We demonstrated that ACME HS simultaneously dissociates and fixes cells, thus preserving morphology and a high RNA integrity number in problematic cell types. This finding renders the ACME HS dissociation method a valuable alternative in scRNA-seq protocols for challenging tissues where obtaining live cell suspension is difficult or impossible.### Competing Interest StatementThe authors have declared no competing interest.

The clinical blood metabogram (CBM) was developed to match a tailored analysis of the blood metabolome to the time, cost, and reproducibility constraints of clinical laboratory testing. By analyzing the main blood metabolite groups, CBM offers clinically relevant information about the intake of low-molecular substances into the organism, humoral regulation, liver function, amino acid level, and the lipid and carbohydrate metabolism. The purpose of this work was to investigate the relevance of using the CBM in patients with diabetes mellitus. For this, a CBM was obtained for 18 healthy individuals, 12 individuals with prediabetes, and 64 individuals with type 2 diabetes mellitus, separated into groups according to fasting blood glucose and oral glucose tolerance tests. The results showed that the CBM reveals diabetes-associated metabolic alterations in the blood, including changes in the levels of carbohydrates, ketone bodies, eicosanoids, phospholipids, and amino acids, which are consistent with the scientific data available to date. The CBM enabled the separation of diabetic patients according to their metabolic metabotypes, providing both a general overview of their metabolic alterations and detailing their individual metabolic characteristics. It was concluded that the CBM is a precise and clinically applicable test for assessing an individual’s metabolic status in diabetes mellitus for diagnostic and treatment purposes.

Searchable abstracts of presentations at key conferences in endocrinology ISSN 1470-3947 (print) | ISSN 1479-6848 (online)

According to the "Federal Law on the Fundamentals of Protection of the Public Health", medical care for patients should be provided in accordance with National Russian guidelines for the relevant nosology, which are based on the principles of evidence-based medicine. The article presents an analysis of the compliance with the completeness of implementation of National Russian guidelines in the treatment of patients with type 2 diabetes mellitus (DM 2) in real clinical practice. The analysis of the actual state of management of DM 2 patients was carried out from the Federal Register of diabetes as of 01.01.2023. Incomplete compliance with the guidelines on the frequency of measuring glycated hemoglobin, the rate of intensification of hypoglycemic therapy, and the appointment of new classes of hypoglycemic drugs was established. Possible reasons for the identified discrepancies between real practice and guidelines requirements are discussed, as well as possible measures to overcome these discrepancies.

Hyperprolactinemia is a persistent excess of the blood serum prolactin. The syndrome contains various symptoms, the most characteristic is a violation of the reproductive system. There are multiple endogenous and exogenous causes of hyperprolactinemia. The main treatment method is dopamine agonist therapy, in case of prolactinoma existence, surgical and radiation methods can be applied. About 15% of patients are resistant to dopamine agonist therapy, which determines creation of individual management tactics. The article presents a draft of clinical guidelines for the diagnosis and treatment of hyperprolactinemia, which provides a modern examination algorithm, discusses the basic principles of diagnostics and treatment approaches.