Mouse vascularized adipose spheroids: an organotypic model for thermogenic adipocytes

Adipose tissues, particularly beige and brown adipose tissue, play crucial roles in energy metabolism. Brown adipose tissues’ thermogenic capacity and the appearance of beige cells within white adipose tissue have spurred interest in their metabolic impact and therapeutic potential. Brown and beige fat cells, activated by environmental factors like cold exposure or by pharmacology, share metabolic mechanisms that drive non-shivering thermogenesis. Understanding these two cell types requires advanced, yet broadly applicable in vitro models that reflect the complex microenvironment and vasculature of adipose tissues. Here we present mouse vascularized adipose spheroids of the stromal vascular microenvironment from inguinal white adipose tissue, a tissue with ‘beiging’ capacity in mice and humans. We show that adding a scaffold improves vascular sprouting, enhances spheroid growth, and upregulates adipogenic markers, thus reflecting increased adipocyte maturity. Transcriptional profiling via RNA sequencing revealed distinct metabolic pathways upregulated in our vascularized adipose spheroids, with increased expression of genes involved in glucose metabolism, lipid metabolism, and thermogenesis. Functional assessment demonstrated increased oxygen consumption in vascularized adipose spheroids compared to classical 2D cultures, which was enhanced by β-adrenergic receptor stimulation correlating with elevated β-adrenergic receptor expression. Moreover, stimulation with the naturally occurring adipokine, FGF21, induced Ucp1 mRNA expression in the vascularized adipose spheroids. In conclusion, vascularized inguinal white adipose tissue spheroids provide a physiologically relevant platform to study how the stromal vascular microenvironment shapes adipocyte responses and influence activated thermogenesis in beige adipocytes

A MAFG-lncRNA axis links systemic nutrient abundance to hepatic glucose metabolism

Obesity and type 2 diabetes mellitus are global emergencies and long noncoding RNAs (lncRNAs) are regulatory transcripts with elusive functions in metabolism. Here we show that a high fraction of lncRNAs, but not protein-coding mRNAs, are repressed during diet-induced obesity (DIO) and refeeding, whilst nutrient deprivation induced lncRNAs in mouse liver. Similarly, lncRNAs are lost in diabetic humans. LncRNA promoter analyses, global cistrome and gain-of-function analyses confirm that increased MAFG signaling during DIO curbs lncRNA expression. Silencing Mafg in mouse hepatocytes and obese mice elicits a fasting-like gene expression profile, improves glucose metabolism, de-represses lncRNAs and impairs mammalian target of rapamycin (mTOR) activation. We find that obesity-repressed LincIRS2 is controlled by MAFG and observe that genetic and RNAi-mediated LincIRS2 loss causes elevated blood glucose, insulin resistance and aberrant glucose output in lean mice. Taken together, we identify a MAFG-lncRNA axis controlling hepatic glucose metabolism in health and metabolic disease

A MAFG-lncRNA axis links systemic nutrient abundance to hepatic glucose metabolism

Obesity and type 2 diabetes mellitus are global emergencies and long noncoding RNAs (lncRNAs) are regulatory transcripts with elusive functions in metabolism. Here we show that a high fraction of lncRNAs, but not protein-coding mRNAs, are repressed during diet-induced obesity (DIO) and refeeding, whilst nutrient deprivation induced lncRNAs in mouse liver. Similarly, lncRNAs are lost in diabetic humans. LncRNA promoter analyses, global cistrome and gain-of-function analyses confirm that increased MAFG signaling during DIO curbs lncRNA expression. Silencing Mafg in mouse hepatocytes and obese mice elicits a fasting-like gene expression profile, improves glucose metabolism, de-represses lncRNAs and impairs mammalian target of rapamycin (mTOR) activation. We find that obesity-repressed LincIRS2 is controlled by MAFG and observe that genetic and RNAi-mediated LincIRS2 loss causes elevated blood glucose, insulin resistance and aberrant glucose output in lean mice. Taken together, we identify a MAFG-lncRNA axis controlling hepatic glucose metabolism in health and metabolic disease

The effects of Rosiglitazone on ovaries of female rats exposed to chemotherapy

Amaç: ÇalıÄ¢mamızda kemoterapötik bir etken madde olan ve ovaryum üzerindeki etkileri geri dönüÄ¢ümsüz olan siklofosfamidin yarattığı ovarial hasarda; PPARG reseptörlerine bağlanarak etkin hale gelen antienflamatuar, anjiyogenik ve immün hücre aktivasyonu üzerindeki baskılayıcı etkileri bilinen Rosiglitazone‟ un etkilerini araÄ¢tırmayı amaçladık. Yöntem: Deney gruplarımız aÄ¢ağıdaki gibi tasarlandı. Kontrol grubu: Hiçbir uygulamaya maruz kalmayan deney grubudur. (n=7) I. Grup Kemoterapi grubu: 100mg/kg siklofosfamid uygulanan deney grubu. Enjeksiyon veya oral herhangi bir tedaviye yönelik ilaç uygulanılmadı, deney grubu diğer gruplarla aynı sürede sakrifiye edildi. (n=7) II. Grup Rosiglitazone deney grubu: 100mg/kg siklofosfamid uygulamasından 3 gün önce ve 15 gün sonra olmak üzere toplamda 18 gün 3mg/kg dozunda rosiglitazone %0.9‟luk NaCl‟ de çözünerek oral gavaj yöntemiyle verildi. Son doz uygulamasından 24 saat sonra deney grubu sakrifiye edildi. (n=7)III. Grup Kemoterapi+NaCl deney grubu: 100mg/kg siklofosfamid uygulamasından 3 gün önce ve 15 gün sonra olmak üzere toplamda 18 gün deney grubuna rosiglitazone eklenmemiÄ¢ %0.9‟lu NaCl solüsyonu oral gavaj yöntemiyle verildi. Son uygulamadan 24 saat sonra deney grubu sakrifiye edildi. (n=7)Deney sonrasında sakrifiye edilen deneklerden alınan ovaryum dokularından alınan kesitler H&E ve Masson‟s Trikrom boyanarak incelendi. Ä mmünohistokimyasal olarak TUNEL boyaması gerçekleÄ¢tirilen kesitlerde TUNEL pozitif boyanan hücrelerin sayılmasıyla DNA fragmantasyonu değerlendirildi. Ovaryum dokularından alınan 4 kesitte folikül sayıları değerlendirildi. Folikül sayıları primordial, preantral ve antral olarak sınıflandırılarak istatistiksel değerlendirmeye alındı. Bulgular: Kontrol grubunun histolojik incelemesinde ovaryum dokusu normal gözlenirken kortekste çok sayıda primordial, primer, sekonder ve olgun foliküller, birkaç korpora lutea ve ovarial siklusa bağlı olarak az sayıda atretik folikül gözlendi. Medullada kan damarları ve gevÄ¢ek bağ doku normal görünümdeydi. Grup 1: Kemoterapi grubunda kortekste yer alan özellikle geliÄ¢mekte olan (primer, sekonder) foliküllerde ve olgun foliküllerde yaygın apoptoz, vakuolizasyon görüldü. Kortekste olgun ve sekonder foliküller, ayrıca korpora lutea çevresinde bağ doku artıÄ¢ı göze çarparken kortikal fibrosisin geliÄ¢tiği alanlar gözlendi. Grup 2: Rosiglitazone deney grubunda yine kemoterapiye bağlı atretik foliküller ve geliÄ¢mekte olan foliküllerde apoptoz birinci ve üçüncü gruba göre daha az oranda görüldü. Kortikal fibrosise bu grupta rastlanmazken foliküller çevresinde de kollajen artıÄ¢ı görülmedi. Grup 3: Kemoterapi + NaCl grubunda birinci gruba benzer Ä¢ekilde apoptoz ve vakuolizasyon yaygındı. Yine kortekste fibrotik alanlar göze çarparken korpora lutea çevresinde kollajen artıÄ¢ı görüldü.TUNEL boyamasında DNA fragmantasyonu değerlendirildiğinde kontrol grubu da dahil olmak üzere apoptoz görüldü. Kontrol grubundaki apoptoz ovaryal siklusun normal süreci olarak kabul edildi. Grup 1 ve Grup 3‟te kontrol grubuna göre TUNEL pozitif hücre sayısı fazla iken bu artıÄ¢ istatistiksel olarak bir anlamlılık ifade etmedi. TUNEL pozitif hücre sayısı Grup 2‟de kontrol grubuna oranla daha az iken yine bu fark istatistiksel olarak anlamlı bir fark değildi. Grup 2 Grup 1 ve Grup 3‟e oranla daha az sayıda TUNEL pozitif hücre içerirken, bu fark sadece Grup 1 ile istatistiksel anlamlılık gösterdi. Kontrol grubu primordial folikül sayısı Grup 1, Grup 2 ve Grup 3‟ten istatistiksel olarak anlamlı ölçüde fazla bulundu. Grup 2‟ye ait primordial folikül sayısı Grup 1 ve Grup 2‟den istatistiksel olarak daha fazlaydı. Grup 1 ile Grup 3 arasında istatistiksel olarak anlamlı bir farklılık görülmedi. Kontrol grubu preantral folikül sayısı Grup 1 ve Grup 3‟ten istatistiksel olarak anlamlı ölçüde fazla bulundu. Kontrol grubu ile Grup 2 preantral folikül sayısı arasında istatistiksel bir farklılık görülmedi. Grup 1 ile Grup 2 ve Grup 3 arasında preantral folikül sayısı açısından bir farklılık bulunmadı. Grup 2 preantral folikül sayısı Grup 3‟ten istatistiksel olarak anlamlı bir Ä¢ekilde fazlaydı. Kontrol grubu antral folikül sayısı Grup 1 ve Grup 3‟ten istatistiksel olarak anlamlı Ä¢ekilde fazla bulundu. Kontrol grubu ile Grup 2 antral folikül sayısında istatistiksel olarak anlamlı bir farklılık görülmedi. Grup 1 ile Grup 2 ve Grup 3 arasında istatistiksel olarak anlamlı bir fark bulunmazken; Grup 2 antral folikül sayısı Grup 3 â€ten istatistiksel olarak anlamlı Ä¢ekilde fazla bulundu. Korpora lutea sayıları arasında anlamlı bir fark tespit edilmedi. Sonuç:ÇalıÄ¢mamızda siklofosfamidin ovaryum üzerindeki apoptotik etkisi hem histolojik hem immünohistokimyasal hem de moleküler tekniklerle, fibrotik etkisi ise ıÄ¢ık mikroskobik olarak gösterilmiÄ¢tir. Rosiglitazone‟un çalıÄ¢mada kullanılan dozuyla siklofosfamidin ovaryum üzerindeki apoptotik etkisini azalttığı gösterilmiÄ¢tir. Ayrıca siklofosfamidin neden olduğu ovarial folikül rezervindeki azalmaya, preantral ve antral foliküllerdeki dejenerasyona karÄ¢ı Rosiglitazone‟un koruyucu bir etkisi olduğu ilk kez ortaya koyulmuÄ¢tur. Ancak Rosiglitazone‟un siklofosfamid toksisitesinden koruyucu etkisinin daha iyi anlaÄ¢ılabilmesi için gelecek çalıÄ¢malarda daha farklı dozçalıÄ¢malarının denenmesi ve foliküller üzerindeki koruyucu etkisinin mekanizmalarının araÄ¢tırılması gerekmektedirPurpose: We aimed to show the effects of Rosiglitazone (which has anti-inflamator, angiogenic and immunosupressive effects) on the irreversible ovarian tissue damage of cyclophosphamide. Material and Method: Animal groups were designed as showed below: Control Group: No application was performed to this animals. (n=7)I. Group Chemotherapy Group: 100 mg/kg cyclophosphamide was applied by intraperitoneal injection. (n=7)II. Group Rosiglitazone Group: 100 mg/kg cyclophosphamide was applied by intraperitoneal injection. Three days before the chemotherapy; 3 mg/kg Rosiglitazone application begun and continiued fifteen consecutive days. Rosiglitazone was applied by orogastric sonda. (n=7)III. Group Chemotherapy + NaCl Group: 100 mg/kg cyclophosphamide was applied by intraperitoneal injection. Three days before the chemotherapy; 1 ml %0.9 NaCl solution application begun and continiued fifteen consecutive days. Rosiglitazone was applied by orogastric sonda. (n=7)At the end of the experiment, animals were sacrified under ether anesthesia. Ovary tissue slides were stained with Hematoxylene&Eosine (H&E) and Masson Trichrome for light microscopy. Immunohistochemical TUNEL stain (terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick-end labeling) was performed to determine apoptosis. The ovary tissues which were collected from the animals. RESULT:In control group; ovary tissue was normal. There were many primordial, primer, seconder, mature follicules and corpora lutea in the ovary cortex. And there was a small number of atretic follicules because of the normal ovarian cycle. Blood vessels and medulla was normal. In Group 1: Chemotherapy group, especially in growing follicules (primer and seconder) and mature follicules there were a widespread apoptosis and vacuolisation. There was cortical fibrosis in cortex and an increase in connective tissue around corpora lutea and mature follicules. There was an increase in atretic follicule numbers. In Group 2: Rosiglitazone group, there were many atretic follicules and apoptosis in growing follicules because of chemotherapy. But the incidances of apoptosis and atretic follicules were lower than Group 1 and 3. Cortical fibrosis and an increase of connective tissue was not seen in this group. In Group 3: Chemotherapy + NaCl group, the findings were similar with Group 1. There were many atretic follicules, widespread apoptosis in growing follicules, cortical fibrosis and increase of connective tissue around the mature follicules and corpora lutea. In TUNEL assay tissues showed widespread TUNEL positive cells even in control group. TUNEL positive cell numbers in control group was evaluated as the normal process of ovarian cycle. There was an increase of TUNEL positive cell counts in Group 1 and Group 3 but this increase was not statistically significant. The rate of TUNEL positive cell numbers was lower in Group 2 tha control group but the difference was not statistically significant. The differences between the Group 2 and Group 3 was found statisticallly significant. Primordial follicule number of control group was found significantly higher than Group 1, Group 2 and Group 3. Number of primordial follicules in Group 2 was found statistically higher than Group 1 and Group 3. There was no significant statistical difference between Group 1 and Group 3.Preantral follicule number of control group was found significantly higher than Group 1 and Group 3. There was no significant difference between Group 2 and Control. The difference between Group 1, Group 2 and Group 3 was not significant statistically. Preantral folicule number of Group 2 was significantly higher than Group 3. Antral follicule number of control group was significantly higher than Group 1 and Group 3. There was no significant difference between control and Group 2. There was not a statistically significancy between Group 1, Group 2 and Group 3. But antral follicule number of Group 2 was significantly higher than Group 3. There was no statistical significancy between corpora lutea numbers of the groups. CONCLUSION: In our study, we showed the fibrotic and apoptotic effect of cyclophosphamide by histologic and immunohistochemical techniques. According to our results; we consider that Rosiglitazone has a diminishing effect on apoptosis caused by cyclophosphamide. Also Rosiglitazone has a diminishing effect on primordial, preantral and antral folicule number reducing effect of cyclophosphamide. In our opinion, more studies are needed to understand the mechanism of Rosiglitazone in chemotherapy damage and if Rosiglitazone has a ability to protect primordial folicule reserve of ovary in chemotherap

lncRNA HOTAIR overexpression induced downregulation of c-Met signaling promotes hybrid epithelial/mesenchymal phenotype in hepatocellular carcinoma cells

Background: Epithelial-to-mesenchymal transition (EMT) and mesenchymal-to-epithelial transition (MET) are both reversible processes, and regulation of phenotypical transition is very important for progression of several cancers including hepatocellular carcinoma (HCC). Recently, it is defined that cancer cells can attain a hybrid epithelial/mesenchymal (hybrid E/M) phenotype. Cells with hybrid E/M phenotype comprise mixed epithelial and mesenchymal properties, they can be more resistant to therapeutics and also more capable of initiating metastatic lesions. However, the mechanisms regulating hybrid E/M in HCC are not well described yet. In this study, we investigated the role of the potential crosstalk between lncRNA HOTAIR and c-Met receptor tyrosine kinase, which are two essential regulators of EMT and MET, in acquiring of hybrid E/M phenotype in HCC