Na(+)-D-glucose cotransporter SGLT1 is pivotal for intestinal glucose absorption and glucose-dependent incretin secretion.

To clarify the physiological role of Na(+)-D-glucose cotransporter SGLT1 in small intestine and kidney, Sglt1(-/-) mice were generated and characterized phenotypically. After gavage of d-glucose, small intestinal glucose absorption across the brush-border membrane (BBM) via SGLT1 and GLUT2 were analyzed. Glucose-induced secretion of insulinotropic hormone (GIP) and glucagon-like peptide 1 (GLP-1) in wild-type and Sglt1(-/-) mice were compared. The impact of SGLT1 on renal glucose handling was investigated by micropuncture studies. It was observed that Sglt1(-/-) mice developed a glucose-galactose malabsorption syndrome but thrive normally when fed a glucose-galactose-free diet. In wild-type mice, passage of D-glucose across the intestinal BBM was predominantly mediated by SGLT1, independent the glucose load. High glucose concentrations increased the amounts of SGLT1 and GLUT2 in the BBM, and SGLT1 was required for upregulation of GLUT2. SGLT1 was located in luminal membranes of cells immunopositive for GIP and GLP-1, and Sglt1(-/-) mice exhibited reduced glucose-triggered GIP and GLP-1 levels. In the kidney, SGLT1 reabsorbed ∼3% of the filtered glucose under normoglycemic conditions. The data indicate that SGLT1 is 1) pivotal for intestinal mass absorption of d-glucose, 2) triggers the glucose-induced secretion of GIP and GLP-1, and 3) triggers the upregulation of GLUT2

Recommendations from the international evidence-based guideline for the assessment and management of polycystic ovary syndrome

Study Question What is the recommended assessment and management of women with polycystic ovary syndrome (PCOS), based on the best available evidence, clinical expertise, and consumer preference? Summary Answer International evidence-based guidelines including 166 recommendations and practice points, addressed prioritized questions to promote consistent, evidence-based care and improve the experience and health outcomes of women with PCOS. What Is Known Already Previous guidelines either lacked rigorous evidence-based processes, did not engage consumer and international multidisciplinary perspectives, or were outdated. Diagnosis of PCOS remains controversial and assessment and management are inconsistent. The needs of women with PCOS are not being adequately met and evidence practice gaps persist. Study Design, Size, Duration International evidence-based guideline development engaged professional societies and consumer organizations with multidisciplinary experts and women with PCOS directly involved at all stages. Appraisal of Guidelines for Research and Evaluation (AGREE) II-compliant processes were followed, with extensive evidence synthesis. The Grading of Recommendations, Assessment, Development, and Evaluation (GRADE) framework was applied across evidence quality, feasibility, acceptability, cost, implementation and ultimately recommendation strength. Participants/Materials, Setting, Methods Governance included a six continent international advisory and a project board, five guideline development groups, and consumer and translation committees. Extensive health professional and consumer engagement informed guideline scope and priorities. Engaged international society-nominated panels included pediatrics, endocrinology, gynecology, primary care, reproductive endocrinology, obstetrics, psychiatry, psychology, dietetics, exercise physiology, public health and other experts, alongside consumers, project management, evidence synthesis, and translation experts. Thirty-seven societies and organizations covering 71 countries engaged in the process. Twenty face-to-face meetings over 15 months addressed 60 prioritized clinical questions involving 40 systematic and 20 narrative reviews. Evidence-based recommendations were developed and approved via consensus voting within the five guideline panels, modified based on international feedback and peer review, with final recommendations approved across all panels. Main Results and the Role of Chance The evidence in the assessment and management of PCOS is generally of low to moderate quality. The guideline provides 31 evidence based recommendations, 59 clinical consensus recommendations and 76 clinical practice points all related to assessment and management of PCOS. Key changes in this guideline include: i) considerable refinement of individual diagnostic criteria with a focus on improving accuracy of diagnosis; ii) reducing unnecessary testing; iii) increasing focus on education, lifestyle modification, emotional wellbeing and quality of life; and iv) emphasizing evidence based medical therapy and cheaper and safer fertility management. Limitations, Reasons for Caution Overall evidence is generally low to moderate quality, requiring significantly greater research in this neglected, yet common condition, especially around refining specific diagnostic features in PCOS. Regional health system variation is acknowledged and a process for guideline and translation resource adaptation is provided. Wider Implications of the Findings The international guideline for the assessment and management of PCOS provides clinicians with clear advice on best practice based on the best available evidence, expert multidisciplinary input and consumer preferences. Research recommendations have been generated and a comprehensive multifaceted dissemination and translation program supports the guideline with an integrated evaluation program. Study Funding/Competing Interest(S) The guideline was primarily funded by the Australian National Health and Medical Research Council of Australia (NHMRC) supported by a partnership with ESHRE and the American Society for Reproductive Medicine. Guideline development group members did not receive payment. Travel expenses were covered by the sponsoring organizations. Disclosures of conflicts of interest were declared at the outset and updated throughout the guideline process, aligned with NHMRC guideline processes. Full details of conflicts declared across the guideline development groups are available at https://www.monash.edu/medicine/sphpm/mchri/pcos/guideline in the Register of disclosures of interest. Of named authors, Dr Costello has declared shares in Virtus Health and past sponsorship from Merck Serono for conference presentations. Prof. Laven declared grants from Ferring, Euroscreen and personal fees from Ferring, Euroscreen, Danone and Titus Healthcare. Prof. Norman has declared a minor shareholder interest in an IVF unit. The remaining authors have no conflicts of interest to declare. The guideline was peer reviewed by special interest groups across our partner and collaborating societies and consumer organizations, was independently assessed against AGREEII criteria and underwent methodological review. This guideline was approved by all members of the guideline development groups and was submitted for final approval by the NHMRC

Recommendations from the 2023 international evidence-based guideline for the assessment and management of polycystic ovary syndrome

Study question What is the recommended assessment and management of those with polycystic ovary syndrome (PCOS), based on the best available evidence, clinical expertise, and consumer preference? Summary answer International evidence-based guidelines address prioritized questions and outcomes and include 254 recommendations and practice points, to promote consistent, evidence-based care and improve the experience and health outcomes in PCOS. What is known already The 2018 International PCOS Guideline was independently evaluated as high quality and integrated multidisciplinary and consumer perspectives from 6 continents; it is now used in 196 countries and is widely cited. It was based on best available, but generally very low- to low-quality, evidence. It applied robust methodological processes and addressed shared priorities. The guideline transitioned from consensus-based to evidence-based diagnostic criteria and enhanced accuracy of diagnosis, whilst promoting consistency of care. However, diagnosis is still delayed, the needs of those with PCOS are not being adequately met, the evidence quality was low, and evidence-practice gaps persist. Study design, size, and duration The 2023 International Evidence-based Guideline update re-engaged the 2018 network across professional societies and consumer organizations with multidisciplinary experts and women with PCOS directly involved at all stages. Extensive evidence synthesis was completed. Appraisal of Guidelines for Research and Evaluation II (AGREEII)-compliant processes were followed. The Grading of Recommendations, Assessment, Development, and Evaluation (GRADE) framework was applied across evidence quality, feasibility, acceptability, cost, implementation, and ultimately recommendation strength, and diversity and inclusion were considered throughout. Participants/materials, setting, and methods This summary should be read in conjunction with the full guideline for detailed participants and methods. Governance included a 6-continent international advisory and management committee, 5 guideline development groups, and paediatric, consumer, and translation committees. Extensive consumer engagement and guideline experts informed the update scope and priorities. Engaged international society-nominated panels included paediatrics, endocrinology, gynaecology, primary care, reproductive endocrinology, obstetrics, psychiatry, psychology, dietetics, exercise physiology, obesity care, public health, and other experts, alongside consumers, project management, evidence synthesis, statisticians, and translation experts. Thirty-nine professional and consumer organizations covering 71 countries engaged in the process. Twenty meetings and 5 face-to-face forums over 12 months addressed 58 prioritized clinical questions involving 52 systematic and 3 narrative reviews. Evidence-based recommendations were developed and approved via consensus across 5 guideline panels, modified based on international feedback and peer review, independently reviewed for methodological rigour, and approved by the Australian Government National Health and Medical Research Council. Main results and the role of chance The evidence in the assessment and management of PCOS has generally improved in the past 5 years but remains of low to moderate quality. The technical evidence report and analyses (∼6000 pages) underpin 77 evidence-based and 54 consensus recommendations, with 123 practice points. Key updates include the following: (1) further refinement of individual diagnostic criteria, a simplified diagnostic algorithm, and inclusion of anti-Müllerian hormone levels as an alternative to ultrasound in adults only; (2) strengthening recognition of broader features of PCOS including metabolic risk factors, cardiovascular disease, sleep apnoea, very high prevalence of psychological features, and high risk status for adverse outcomes during pregnancy; (3) emphasizing the poorly recognized, diverse burden of disease and the need for greater healthcare professional education, evidence-based patient information, improved models of care, and shared decision-making to improve patient experience, alongside greater research; (4) maintained emphasis on healthy lifestyle, emotional well-being, and quality of life, with awareness and consideration of weight stigma; and (5) emphasizing evidence-based medical therapy and cheaper and safer fertility management. Limitations and reasons for caution Overall, recommendations are strengthened and evidence is improved but remains generally low to moderate quality. Significantly greater research is now needed in this neglected, yet common condition. Regional health system variation was considered and acknowledged, with a further process for guideline and translation resource adaptation provided. Wider implications of the findings The 2023 International Guideline for the Assessment and Management of PCOS provides clinicians and patients with clear advice on best practice, based on the best available evidence, expert multidisciplinary input, and consumer preferences. Research recommendations have been generated, and a comprehensive multifaceted dissemination and translation programme supports the guideline with an integrated evaluation programme

Recommendations from the 2023 international evidence-based guideline for the assessment and management of polycystic ovary syndrome

STUDY QUESTION What is the recommended assessment and management of those with polycystic ovary syndrome (PCOS), based on the best available evidence, clinical expertise, and consumer preference? SUMMARY ANSWER International evidence-based guidelines address prioritized questions and outcomes and include 254 recommendations and practice points, to promote consistent, evidence-based care and improve the experience and health outcomes in PCOS. WHAT IS KNOWN ALREADY The 2018 International PCOS Guideline was independently evaluated as high quality and integrated multidisciplinary and consumer perspectives from six continents; it is now used in 196 countries and is widely cited. It was based on best available, but generally very low to low quality, evidence. It applied robust methodological processes and addressed shared priorities. The guideline transitioned from consensus based to evidence-based diagnostic criteria and enhanced accuracy of diagnosis, whilst promoting consistency of care. However, diagnosis is still delayed, the needs of those with PCOS are not being adequately met, evidence quality was low and evidence-practice gaps persist. STUDY DESIGN, SIZE, DURATION The 2023 International Evidence-based Guideline update reengaged the 2018 network across professional societies and consumer organizations, with multidisciplinary experts and women with PCOS directly involved at all stages. Extensive evidence synthesis was completed. Appraisal of Guidelines for Research and Evaluation-II (AGREEII)-compliant processes were followed. The Grading of Recommendations, Assessment, Development, and Evaluation (GRADE) framework was applied across evidence quality, feasibility, acceptability, cost, implementation and ultimately recommendation strength and diversity and inclusion were considered throughout. PARTICIPANTS/MATERIALS, SETTING, METHODS This summary should be read in conjunction with the full Guideline for detailed participants and methods. Governance included a six-continent international advisory and management committee, five guideline development groups, and paediatric, consumer, and translation committees. Extensive consumer engagement and guideline experts informed the update scope and priorities. Engaged international society-nominated panels included paediatrics, endocrinology, gynaecology, primary care, reproductive endocrinology, obstetrics, psychiatry, psychology, dietetics, exercise physiology, obesity care, public health and other experts, alongside consumers, project management, evidence synthesis, statisticians and translation experts. Thirty-nine professional and consumer organizations covering 71 countries engaged in the process. Twenty meetings and five face-to-face forums over 12 months addressed 58 prioritized clinical questions involving 52 systematic and 3 narrative reviews. Evidence-based recommendations were developed and approved via consensus across five guideline panels, modified based on international feedback and peer review, independently reviewed for methodological rigour, and approved by the Australian Government National Health and Medical Research Council (NHMRC). MAIN RESULTS AND THE ROLE OF CHANCE The evidence in the assessment and management of PCOS has generally improved in the past five years, but remains of low to moderate quality. The technical evidence report and analyses (∼6000 pages) underpins 77 evidence-based and 54 consensus recommendations, with 123 practice points. Key updates include: i) further refinement of individual diagnostic criteria, a simplified diagnostic algorithm and inclusion of anti-Müllerian hormone (AMH) levels as an alternative to ultrasound in adults only; ii) strengthening recognition of broader features of PCOS including metabolic risk factors, cardiovascular disease, sleep apnea, very high prevalence of psychological features, and high risk status for adverse outcomes during pregnancy; iii) emphasizing the poorly recognized, diverse burden of disease and the need for greater healthcare professional education, evidence-based patient information, improved models of care and shared decision making to improve patient experience, alongside greater research; iv) maintained emphasis on healthy lifestyle, emotional wellbeing and quality of life, with awareness and consideration of weight stigma; and v) emphasizing evidence-based medical therapy and cheaper and safer fertility management. LIMITATIONS, REASONS FOR CAUTION Overall, recommendations are strengthened and evidence is improved, but remains generally low to moderate quality. Significantly greater research is now needed in this neglected, yet common condition. Regional health system variation was considered and acknowledged, with a further process for guideline and translation resource adaptation provided. WIDER IMPLICATIONS OF THE FINDINGS The 2023 International Guideline for the Assessment and Management of PCOS provides clinicians and patients with clear advice on best practice, based on the best available evidence, expert multidisciplinary input and consumer preferences. Research recommendations have been generated and a comprehensive multifaceted dissemination and translation program supports the Guideline with an integrated evaluation program. STUDY FUNDING/COMPETING INTEREST(S) This effort was primarily funded by the Australian Government via the National Health Medical Research Council (NHMRC) (APP1171592), supported by a partnership with American Society for Reproductive Medicine, Endocrine Society, European Society for Human Reproduction and Embryology, and European Society for Endocrinology. The Commonwealth Government of Australia also supported Guideline translation through the Medical Research Future Fund (MRFCRI000266). HJT and AM are funded by NHMRC fellowships. JT is funded by a Royal Australasian College of Physicians (RACP) fellowship. Guideline development group members were volunteers. Travel expenses were covered by the partnering organizations. Disclosures of interest were strictly managed according to NHMRC policy and are available with the full guideline, technical evidence report, peer review and responses (www.monash.edu/medicine/mchri/pcos). Of named authors HJT, CTT, AD, LM, LR, JBoyle, AM have no conflicts of interest to declare. JL declares grant from Ferring and Merck; consulting fees from Ferring and Titus Health Care; speaker’s fees from Ferring; unpaid consultancy for Ferring, Roche Diagnostics and Ansh Labs; and sits on advisory boards for Ferring, Roche Diagnostics, Ansh Labs, and Gedeon Richter. TP declares a grant from Roche; consulting fees from Gedeon Richter and Organon; speaker’s fees from Gedeon Richter and Exeltis; travel support from Gedeon Richter and Exeltis; unpaid consultancy for Roche Diagnostics; and sits on advisory boards for Roche Diagnostics. MC declares travels support from Merck; and sits on an advisory board for Merck. JBoivin declares grants from Merck Serono Ltd.; consulting fees from Ferring B.V; speaker’s fees from Ferring Arzneimittell GmbH; travel support from Organon; and sits on an advisory board for the Office of Health Economics. RJN has received speaker’s fees from Merck and sits on an advisory board for Ferring. AJoham has received speaker’s fees from Novo Nordisk and Boehringer Ingelheim. The guideline was peer reviewed by special interest groups across our 39 partner and collaborating organizations, was independently methodologically assessed against AGREEII criteria and was approved by all members of the guideline development groups and by the NHMRC

Optimalne metode otkrivanja antigena za prijenosnike organskih aniona u kriostatskim narescima štakorskog bubrega

To localise antigens by immunocytochemistry (IC), the samples of tissues or cells are usually denatured by fi xation, and either frozen and cryosectioned, or embedded in paraffi n before sectioning. p-Formaldehyde (PFA; formalin) is a common fi xative, which preserves antigenicity of proteins, but damages the tissue/cell morphology and “masks” the antibody binding sites (epitopes). In order to “unmask” epitopes, some kind of antigen retrieval (AR) is used. The aim of this study was: a) to fi nd an optimal AR method in cryosections of in vivo PFA-fi xed kidneys for organic anion transporters (Oat) that reside in the basolateral (Oat1, Oat3) and brush-border membrane (Oat2, Oat5) of the rat renal proximal tubules, and b) using optimal method, to compare IC staining of Oats in kidneys that had been PFA-fi xed in vivo or in vitro. IC staining in untreated cryosections was compared with that following detergent treatment or microwave heating in citrate buffer of pH 3, pH 6, or pH 8, with or without alcohol pre-treatment. The preferred AR method for Oat1, Oat2, and Oat5 was heating of cryosections at pH 6, and for Oat3 heating at pH 3, without alcohol pre-treatment. Compared with tissue fi xed in vivo, tissue fi xed in vitro exhibited damaged tubule morphology, similar staining intensity of Oat1 and Oat3, and higher staining intensity of Oat2 and Oat5. We conclude that for optimal IC presentation, each Oat in the rat kidney has to be treated individually, with different fi xation and AR approach.Za imunocitokemijsku (IC) lokalizaciju antigena uzorci tkiva ili stanica obično se denaturiraju fi ksacijom, a potom se odmah smrznu i naresci režu kriostatom ili se uklope u parafi n prije rezanja narezaka. Parafi n se kasnije odstrani alkoholima. p-Formaldehid (PFA; formalin) čest je fi ksativ, koji čuva antigeničnost proteina, ali oštećuje morfologiju stanica i “maskira” vezna mjesta za protutijela (epitope). Za “demaskiranje” epitopa potrebno je primijeniti neku od metoda otkrivanja (regeneracije) antigena (RA). Namjera ove studije jest: a) odrediti optimalnu metodu RA u kriostatskim narescima PFA-fi ksiranih bubrega in vivo za prijenosnike organskih aniona (Oat) smještene u bazolateralnoj (Oat1, Oat3) i četkastoj (Oat2, Oat5) membrani proksimalnih kanalića u bubrezima štakora i b) rabeći optimalnu metodu, usporediti IC bojenje Oat-a u bubrezima koji su bili PFA-fi ksirani in vivo ili in vitro. IC bojenje u neobrađenim kriostatskim narescima uspoređeno je s bojenjem nakon obrade detergentom ili mikrovalnim kuhanjem u citratnom puferu pH 3, pH 6 ili pH 8, s prethodnom obradom alkoholima ili bez nje. Optimalni RA-uvjet za Oat1, Oat2 i Oat5 bio je kuhanje narezaka pri pH 6, a za Oat3 pri pH 3, bez prethodne obrade alkoholima. U usporedbi s tkivom fi ksiranim in vivo, tkivo fi ksirano in vitro imalo je: oštećenu morfologiju kanalića, sličnu jačinu bojenja Oat1 i Oat3, a jače bojenje Oat2 i Oat5. Zaključujemo da svaki Oat u bubregu štakora treba obraditi pojedinačno, s različitom fi ksacijom i metodom RA, kako bi se našli optimalni uvjeti za njegovo IC prikazivanje

Renal expression of organic anion transporter Oat5 in rats and mice exhibits the female-dominant sex differences

The organic anion transporter 5 (Oat5, Slc22a19) was previously localized to the brush-border of proximal tubule (PT) S3 segment in rat and mouse kidneys. Here we report on sex hormone-regulated expression of Oat5 in rat kidneys, after reinvestigating: a) expression of its mRNA by end-point and real time RT-PCR in the tissue, b) abundance of its protein by Western blotting (WB) in isolated membranes, and c) immunolocalization in tissue cryosections. In untreated male (M) and female (F) adult rats, the expression of Oat5 mRNA was predominant in the outer stripe (OS), exhibiting sex differences (M<F), upregulated by castration, and unaffected by ovariectomy. In castrated M, testosterone treatment strongly downregulated, whereas estradiol and progesterone treatment weakly upregulated its expression. By WB, a single protein band of ~72 kDa in variously-treated animals exhibited a density pattern comparable to that of mRNA. By immunostaining, Oat5 protein was localized to the brush-border of S1/S2 in the cortex (CO) (weakly) and in S3 of the OS and medullary rays (strongly) with the F-dominant intensity. In variously-treated rats, the immunostaining pattern matched that of mRNA and WB data. In prepubertal rats, the renal expression of Oat5 mRNA and protein was weak and sex-independent. In adult mice, the sex-dependent pattern of renal Oat5 protein expression was comparable to that in rats. Therefore, the renal expression of Oat5 in rats (and mice) exhibits zonal (CO<OS) and sex differences (M<F), which appear after puberty, largely due to androgen-driven downregulation of its mRNA and protein expression