Role of KLF4 and SIAT7A interaction accelerates myocardial hypertrophy induced by Ang II.

Sialylation catalysed by sialyltransferase 7A (SIAT7A) plays a role in the development of cardiac hypertrophy. However, the regulatory mechanisms upstream of SIAT7A in this context remain poorly elucidated. Previous study demonstrated that KLF4 activates the SIAT7A gene in ischemic myocardium by binding to its promoter region. Nevertheless, the potential involvement of KLF4 in regulating SIAT7A expression in Ang II-induced hypertrophic cardiomyocytes remains uncertain. This study seeks to deepen the underlying mechanisms of the KLF4 and SIAT7A interaction in the progression of Ang II-induced cardiac hypertrophy. The results showed a concurrent increase in SIAT7A and KLF4 levels in hypertrophic myocardium of essential hypertension patients and in hypertrophic cardiomyocytes stimulated by Ang II. In vitro experiments revealed that reducing KLF4 levels led to a decrease in both SIAT7A synthesis and Sialyl-Tn antigen expression, consequently inhibiting Ang II-induced cardiomyocyte hypertrophy. Intriguingly, reducing SIAT7A levels also resulted in decreased KLF4 expression and suppression cardiomyocyte hypertrophy. Consistent with this, elevating SIAT7A levels increased KLF4 expression and exacerbated cardiomyocyte hypertrophy in both in vivo and in vitro experiments. Additionally, a time-course analysis indicated that KLF4 expression preceded that of SIAT7A. Luciferase reporter assays further confirmed that modulating SIAT7A levels directly influenced the transcriptional activity of KLF4 in cardiomyocytes. In summary, KLF4 expression is upregulated in cardiomyocytes treated with Ang II, which subsequently induces the expression of SIAT7A. The elevated levels of SIAT7A, in turn, enhance the transcription of KLF4. These findings suggest a positive feedback loop between KLF4 and SIAT7A-Sialyl-Tn, ultimately promoting Ang II-induced cardiac hypertrophy.

Mechanisms of epigenomic and functional convergence between glucocorticoid- and IL4-driven macrophage programming.

Macrophages adopt distinct phenotypes in response to environmental cues, with type-2 cytokine interleukin-4 promoting a tissue-repair homeostatic state (M2IL4). Glucocorticoids (GC), widely used anti-inflammatory therapeutics, reportedly impart a similar phenotype (M2GC), but how such disparate pathways may functionally converge is unknown. We show using integrative functional genomics that M2IL4 and M2GC transcriptomes share a striking overlap mirrored by a shift in chromatin landscape in both common and signal-specific gene subsets. This core homeostatic program is enacted by transcriptional effectors KLF4 and the glucocorticoid receptor, whose genome-wide occupancy and actions are integrated in a stimulus-specific manner by the nuclear receptor cofactor GRIP1. Indeed, many of the M2IL4:M2GC-shared transcriptomic changes were GRIP1-dependent. Consistently, GRIP1 loss attenuated phagocytic activity of both populations in vitro and macrophage tissue-repair properties in the murine colitis model in vivo. These findings provide a mechanistic framework for homeostatic macrophage programming by distinct signals, to better inform anti-inflammatory drug design.

Transcriptomic Profiling of Primary Microglia: Effects of miR-19a-3p and miR-19b-3p on Microglia Activation.

Neuropathic pain resulting from spinal cord injury (SCI) is a significant secondary health issue affecting around 60% of individuals with SCI. After SCI, activation of microglia, the immune cells within the central nervous system, leads to neuroinflammation by producing pro-inflammatory cytokines and affects neuropathic pain. This interplay between inflammation and pain contributes to the persistent and intense pain experienced by many individuals with SCI. MicroRNAs (miRs) have been critical regulators of neuroinflammation. Previous research in our laboratory has revealed upregulation levels of circulating miR-19a and miR-19b in individuals with SCI with neuropathic pain compared to those without pain. In this study, we treated primary microglial cultures from mice with miR-19a and miR-19b for 24 h and conducted RNA sequencing analysis. Our results showed that miR-19a and miR-19b up- and downregulate different genes according to the volcano plots and the heatmaps. miR-19a and miR-19b regulate inflammation through distinct signaling pathways. The results showed that miR-19a promotes inflammation via toll-like receptor signaling, TNF signaling, and cytokine-cytokine receptor interactions, while miR-19b increases inflammatory responses through the PI3K-Akt signaling pathway, focal adhesion, and extracellular matrix receptor interactions. The protein-protein interaction (PPI) networks used the STRING database to identify transcription factors associated with genes up- or downregulated by miR-19a and miR-19b. Key transcription factors, such as STAT1, STAT2, and KLF4 for miR-19a, and Nr4a1, Nr4a2, and Nr4a3 for miR-19b, were identified and revealed their roles in regulating neuroinflammation. This study demonstrates that miR-19a and miR-19b modulate diverse patterns of gene expression, regulate inflammation, and induce inflammatory responses in microglia.

MicroRNA Inhibiting Atheroprotective Proteins in Patients with Unstable Angina Comparing to Chronic Coronary Syndrome.

Patients with unstable angina present clinical characteristics of atherosclerotic plaque vulnerability, contrary to chronic coronary syndrome patients. The process of athersclerotic plaque destabilization is also regulated by microRNA particles. In this study, the investigation on expression levels of microRNAs inhibiting the expression of proteins that protect from atherosclerotic plaque progression (miR-92a inhibiting KLF2, miR-10b inhibiting KLF4, miR-126 inhibiting MerTK, miR-98 inhibiting IL-10, miR-29b inhibiting TGFß1) was undertaken. A number of 62 individuals were enrolled-unstable angina (UA, n = 14), chronic coronary syndrome (CCS, n = 38), and healthy volunteers (HV, n = 10). Plasma samples were taken, and microRNAs expression levels were assessed by qRT-PCR. As a result, the UA patients presented significantly increased miR-10b levels compared to CCS patients (0.097 vs. 0.058, p = 0.033). Moreover, in additional analysis when UA patients were grouped together with stable patients with significant plaque in left main or proximal left anterior descending ("UA and LM/proxLAD" group, n = 29 patients) and compared to CCS patients with atherosclerotic lesions in other regions of coronary circulation ("CCS other" group, n = 25 patients) the expression levels of both miR-10b (0.104 vs. 0.046; p = 0.0032) and miR-92a (92.64 vs. 54.74; p = 0.0129) were significantly elevated. In conclusion, the study revealed significantly increased expression levels of miR-10b and miR-92a, a regulator of endothelial protective KLF factors (KLF4 and KLF2, respectively) in patients with more vulnerable plaque phenotypes.

miR-7 promotes apoptosis and autophagy of granulosa cells by targeting KLF4 via JAK/STAT3 signaling pathway in chickens.

Granulosa cell (GC) death, which leads to follicular atresia, primarily occurs through apoptosis and autophagy. miRNAs are known to be key regulators of autophagy and apoptosis. Although miR-7 acting as a key regulator of follicular atresia, its precise role in granulosa cell autophagy and apoptosis remains to be fully elucidated. In this study, we found that miR-7 was highly expressed in the follicle based on qPCR analysis. Subsequently, transfection of miR-7 inhibitors and mimics downregulated or upregulated the expression of miR-7 and promoted autophagic and apoptotic processes in chicken follicle granulosa cells. Mechanistically, through dual-luciferase reporter gene assays, we validated that KLF4 is a target gene of miR-7. Contrarily, KLF4 was found to negatively regulate autophagy and apoptosis in follicular granulosa cells as evidenced by genetic intervention of KLF4 silencing and overexpression. Furthermore, JAK/STAT3 signaling pathway was confirmed to mediate the regulation of miR-7-KLF4 axis on GC autophagy and apoptosis. These findings offer evidences of the crucial involvement of the miR-7-KLF4 signaling axis in determining autophagy and apoptosis of GCs. This study could offer an important theoretical basis for the use of molecular-assisted breeding in chickens.

Low Intraocular Pressure Induces Fibrotic Changes in the Trabecular Meshwork and Schlemm's Canal of Sprague Dawley Rats.

Purpose: Continuous artificial aqueous humor drainage in the eyes of patients with glaucoma undergoing trabeculectomy likely exerts abnormal shear stress. However, it remains unknown how changes in intraocular pressure (IOP) can affect aqueous humor outflow (AHO). Methods: Here, we induced and maintained low intraocular pressure (L-IOP) in healthy Sprague Dawley (SD) rats by puncturing their eyes using a tube (200-µm diameter) for 2 weeks. After the rats were euthanized, their eyes were removed, fixed, embedded, stained, and scanned to analyze the physiological and pathological changes in the trabecular meshwork (TM) and Schlemm's canal (SC). We measured SC parameters using ImageJ software and assessed the expression of various markers related to flow shear stress (KLF4), fibrosis (TGF-ß1, TGF-ß2, α-SMA, pSmad1/5, pSmad2/3, and fibronectin), cytoskeleton (integrin ß1 and F-actin), diastolic function (nitric oxide synthase and endothelial nitric oxide synthase [eNOS]), apoptosis (cleaved caspase-3), and proliferation (Ki-67) using immunofluorescence or immunohistochemistry. Results: L-IOP eyes showed a larger SC area, higher eNOS expression, and lower KLF4 and F-actin expression in the TM and SC (both P < 0.05) than control eyes. The aqueous humor of L-IOP eyes had a higher abundance of fibrotic proteins and apoptotic cells than that of control eyes, with significantly higher TGF-ß1, α-SMA, fibronectin, and cleaved caspase-3 expression (all P < 0.05). Conclusions: In conclusion, a persistence of L-IOP for 2 weeks may contribute to fibrosis in the TM and SC and might be detrimental to conventional AHO in SD rat eyes. Translational Relevance: Clinicians should consider that aberrant shear force induced by aqueous humor fluctuation may damage AHO outflow channel when treating patients.

Low shear stress protects chondrocytes from IL-1ß-induced apoptosis by activating ERK5/KLF4 signaling and negatively regulating miR-143-3p.

OBJECTIVE: This study investigated the protective effects of low fluid shear stress (FSS ≤ 2 dyn/cm²) against interleukin-1ß (IL-1ß)-induced chondrocyte apoptosis and explored the underlying molecular mechanisms. METHODS: Chondrocytes were cultured under four conditions: control, IL-1ß stimulation, low FSS, and combined low FSS + IL-1ß stimulation. Apoptosis was assessed using Hoechst staining and flow cytometry. Western blotting determined the expression of caspase-3 (CASP3), caspase-8 (CASP8), and NF-κB p65. Quantitative real-time PCR measured miR-143-3p expression. The roles of miR-143-3p and the extracellular signal-regulated kinase 5 (ERK5)/Krüppel-like factor 4 (KLF4) signaling pathway were further investigated using miR-143-3p mimics and inhibitors, an ERK5 inhibitor, and a KLF4 overexpression vector. RESULTS: IL-1ß induced significant chondrocyte apoptosis, which was markedly inhibited by low FSS. Mechanistically, low FSS suppressed miR-143-3p expression, thereby enhancing ERK5 signaling. This activated ERK5 subsequently upregulated KLF4 expression, further mitigating IL-1ß-induced damage. Importantly, miR-143-3p overexpression under low FSS conditions exacerbated IL-1ß-induced apoptosis, while miR-143-3p inhibition attenuated it. Consistent with this, ERK5 inhibition augmented IL-1ß-induced apoptosis, whereas KLF4 overexpression suppressed it. CONCLUSION: Low FSS protects chondrocytes from IL-1ß-induced apoptosis by suppressing miR-143-3p and activating the ERK5/KLF4 signaling pathway. This study reveals a novel mechanism by which mechanical stimulation protects cartilage.

KLF4 regulates trophoblast function and associates with unexplained recurrent spontaneous abortion.

BACKGROUND: Recurrent spontaneous abortion (RSA) is defined as two or more consecutive spontaneous abortions before 20 weeks with the same spouse [1]. However, approximately 50% of RSA cases of unknown cause are classified as unexplained recurrent spontaneous abortion (URSA). Potential factors include decreased trophoblast cell migration and invasion, leading to impaired placental implantation and maintenance of the normal maternal-fetal interface. However, the mechanism of this pathogenesis remains unknown. In this study, we investigated the potential role and mechanism of KLF4 in regulating URSA by influencing the invasion and migration ability of trophoblast cells. METHODS: We firstly identified 817 differentially expressed genes by performing a difference analysis of the dataset GSE121950 [2] related to recurrent abortion, and intersected the top 10 genes obtained respectively by the three algorithms: DMNC, MNC, and EPC using Venn Diagram.To detect the expression levels of core genes, villi samples were obtained from normal pregnant women and patients with URSA. RT-qPCR analysis revealed a significant difference in KLF4 mRNA expression and KLF4 was then analyzed. Trophoblast cell lines HTR8 and JEG3 were used to investigate the effect of KLF4 on trophoblastic function. Wound healing and transwell assays was performed to detect the invasion and migration of trophoblast cells. The expression of epithelial-mesenchymal transition(EMT) molecules were detected by RT-qPCR and western blot. Promoter detection and epigenetic modification were detected by chromatin immunoprecipitation (ChIP) assay. Molecular nuclear localization was detected by immunofluorescence and subcellular fractionation. Miscarried mice model was used to study the effects of KLF4 on URSA induced by reduced trophoblast invasion and migration. RESULTS: KLF4 is highly expressed in the villi of patients with URSA. KLF4 inhibits the expression level of H3R2ME2a in trophoblast cells by regulating the transcriptional level and nuclear translocation of PRMT6, thereby inhibiting the possible regulatory mechanism of trophoblastic invasion and providing a potential treatment strategy for URSA in vivo. CONCLUSIONS: The KLF4/PRMT6/H3R2ME2a axis regulates mechanisms associated with unexplained recurrent spontaneous abortion by regulating trophoblast function.

A Sendai virus-based expression system directs efficient induction of chondrocytes by transcription factor-mediated reprogramming.

Cartilage rarely heals spontaneously once damaged. Osteoarthritis (OA) is the most common degenerative joint disease among the elderly; however, effective treatment for OA is currently lacking. Autologous chondrocyte implantation (ACI), an innovative regenerative technology involving the implantation of healthy chondrocytes, may restore damaged lesions. Chondrocytes for ACI may potentially be induced from differentiated somatic cells using retrovirus (RV)-mediated transduction of three reprogramming factors (SOX9, KLF4, and c-MYC). However, the efficiency of the current induction system needs to be improved and the safety issues arising from the genomic integration of the vector DNA have to be addressed. To solve these problems, we used an RNA vector, termed the replication-defective and persistent Sendai virus vector (SeVdp), to express reprogramming factors for chondrocyte induction. Our results showed that the SeVdp-based vector induced chondrocytes more efficiently than the RV vector, probably because of robust and rapid expression of the transgenes, without any apparent integration of the SeVdp vector. The induced chondrocytes formed cartilage-like tissues when injected subcutaneously into mice. Thus, the SeVdp-based system for inducing chondrocytes may act as a foundation for developing safer and more effective treatments for damaged cartilage.

Repressor elements provide insights into tissue development and phenotypes in pigs.

Repressor elements significantly influence economically relevant phenotypes in pigs; however, their precise roles and characteristics are inadequately understood. In the present study, we employed H3K27me3 profiling, assay for transposase-accessible chromatin with highthroughput sequencing (ATAC-seq), and RNA sequencing (RNA-seq) data across six tissues derived from three embryonic layers to identify and map 2 034 super repressor elements (SREs) and 22 223 typical repressor elements (TREs) in the pig genome. Notably, many repressor elements were conserved across mesodermal and ectodermal tissues. SREs exhibited tight regulation of their target genes, affecting a limited number of genes within a specific genomic region with pronounced effects, while TREs exerted broader but weaker regulation over a wider range of target genes. Furthermore, in neuronal tissues, genes regulated by repressor elements started to be repressed during the differentiation of stem cells into progenitor cells. Notably, analysis showed that many repressor elements exhibited cooperative and additive effects on the modulation of KLF4 expression. This research provides the first comprehensive map of pig repressor elements, serving as an essential reference for future studies on repressor elements.

Circular RNA hsa_circ_0008726 Targets the hsa-miR-206-3p/KLF4 Axis to Modulate 4,4'-Methylene Diphenyl Diisocyanate-Glutathione Conjugate-Induced Chemokine Transcription in Macrophages.

Exposure to 4,4'-methylene diphenyl diisocyanate (MDI) in the workplace may lead to the development of occupational asthma (OA). However, the specific mechanism(s) by which MDI induces OA are poorly understood. Previous reports have demonstrated that MDI and MDI-glutathione (GSH) conjugate exposure downregulates endogenous human/murine (hsa/mmu)-microRNA(miR)-206-3p, resulting in the activation of mmu/hsa-miR-206-3p-regulated signaling pathways in macrophages. Circular RNAs (circRNAs) regulate many important biological processes by targeting endogenous miRs; however, whether MDI/MDI-GSH exposure may influence circRNA expressions is unknown. Several circRNAs have been identified that regulate hsa-miR-206-3p. We hypothesize that MDI-GSH conjugate exposure induces endogenous circRNA(s) to regulate hsa-miR-206-3p in macrophages. The expression of candidate hsa-miR-206-3p-binding circRNAs was determined from MDI-GSH conjugate-treated differentiated THP-1 macrophages using RT-qPCR. MDI-GSH exposures induced hsa_circ_0008726 and its host gene transcript DNAJB6, whereas other circRNA(s) examined were either not detected or unchanged. RNA-induced silencing complex-immunoprecipitation (RISC-IP) experiments confirm that hsa-miR-206-3p can bind to hsa_circ_0008726. The expressions of endogenous hsa-miR-206-3p, hsa-miR-206-3p-regulated KLF4, and KLF4-activated M2 macrophage-associated markers and chemokines were up-/down-regulated by transfection of hsa_circ_0008726 siRNAs or hsa_circ_0008726 overexpression plasmid in macrophages, respectively. These results suggest MDI-GSH exposure downregulates hsa-miR-206-3p via induction of endogenous hsa_circ_0008726/DNAJB6, resulting in the upregulation of hsa-miR-206-3p-mediated regulations in macrophages.

Berberine Inhibits KLF4 Promoter Methylation and Ferroptosis to Ameliorate Diabetic Nephropathy in Mice.

Inflammation, oxidative stress, fibrosis, and ferroptosis play important roles in diabetic nephropathy development. Krüppel-like factor 4 (KLF4) is a transcriptional factor, which regulates multiple cell processes and is involved in diabetic nephropathy. Berberine has various biological activities, including anti-inflammation, antioxidative stress, and antiferroptosis. Berberine has been shown to inhibit diabetic nephropathy, but whether it involves KLF4 and ferroptosis remains unknown. We established a diabetic nephropathy mice model and administered berberine to the mice. The kidney function, renal structure and fibrosis, expression of KLF4 and DNA methylation enzymes, DNA methylation of the KLF4 promoter, mitochondria structure, and expression of oxidative stress and ferroptosis markers were analyzed. Berberine rescued kidney function and renal structure and prevented renal fibrosis in diabetic nephropathy mice. Berberine suppressed the expression of DNMT1 and DNMT2 and upregulated KLF4 expression by preventing KLF4 promoter methylation. Berberine inhibited the expression of oxidative stress and ferroptosis markers, maintained mitochondria structure, and prevented ferroptosis. Berberine ameliorates diabetic nephropathy by inhibiting Klf4 promoter methylation and ferroptosis.

A suboptimal OCT4-SOX2 binding site facilitates the naïve-state specific function of a Klf4 enhancer.

Enhancers have critical functions in the precise, spatiotemporal control of transcription during development. It is thought that enhancer grammar, or the characteristics and arrangements of transcription factor binding sites, underlie the specific functions of developmental enhancers. In this study, we sought to identify grammatical constraints that direct enhancer activity in the naïve state of pluripotency, focusing on the enhancers for the naïve-state specific gene, Klf4. Using a combination of biochemical tests, reporter assays, and endogenous mutations in mouse embryonic stem cells, we have studied the binding sites for the transcription factors OCT4 and SOX2. We have found that the three Klf4 enhancers contain suboptimal OCT4-SOX2 composite binding sites. Substitution with a high-affinity OCT4-SOX2 binding site in Klf4 enhancer E2 rescued enhancer function and Klf4 expression upon loss of the ESRRB and STAT3 binding sites. We also observed that the low-affinity of the OCT4-SOX2 binding site is crucial to drive the naïve-state specific activities of Klf4 enhancer E2. Altogether, our work suggests that the affinity of OCT4-SOX2 binding sites could facilitate enhancer functions in specific states of pluripotency.

Effect of Kruppel-like factor 4 on PTZ-induced acute seizure mice.

Kruppel-like factor 4 (Klf4) is a transcription factor that is involved in neuronal regeneration and the development of glutamatergic systems. However, it is unknown whether Klf4 is involved in acute seizure. To investigate the potential role of Klf4 in pentylenetetrazol (PTZ)-induced seizure, western blotting, immunofluorescence, behaviour test and electrophysiology were conducted in this study. We found that Klf4 protein and mRNA expression were increased in both the hippocampus (HP) and prefrontal cortex (PFC) after PTZ-induced seizure in mice. HP-specific knockout (KO) of Klf4 in mice decreased protein expression of Klf4 and the down-stream Klf4 target tumour protein 53 (TP53/P53). These molecular changes are accompanied by increased seizure latency, reduced immobility time in the forced swimming test and tail suspension test. Reduced hippocampal protein levels for synaptic proteins, including glutamate receptor 1 (GRIA1/GLUA1) and postsynaptic density protein 95 (DLG4/PSD95), were also observed after Klf4-KO, while increased mRNA levels of complement proteins were observed for complement component 1q subcomponent A (C1qa), complement component 1q subcomponent B (C1qb), complement component 1q subcomponent C (C1qc), complement component 3 (C3), complement component 4A (C4a) and complement component 4B (C4b). Moreover, c-Fos expression induced by PTZ was reduced by hippocampal conditional KO of Klf4. Electrophysiology showed that PTZ-induced action potential frequency was decreased by overexpression of Klf4. In conclusion, these findings suggest that Klf4 plays an important role in regulating PTZ-induced seizures and therefore constitutes a new molecular target that should be explored for the development of antiepileptic drugs.

Targeted partial reprogramming of age-associated cell states improves markers of health in mouse models of aging.

Aging is a complex multifactorial process associated with epigenome dysregulation, increased cellular senescence, and decreased rejuvenation capacity. Short-term cyclic expression of octamer-binding transcription factor 4 (Oct4), sex-determining region Y-box 2 (Sox2), Kruppel-like factor 4 (Klf4), and cellular myelocytomatosis oncogene (cMyc) (OSKM) in wild-type mice improves health but fails to distinguish cell states, posing risks to healthy cells. Here, we delivered a single dose of adeno-associated viruses (AAVs) harboring OSK under the control of the cyclin-dependent kinase inhibitor 2a (Cdkn2a) promoter to specifically partially reprogram aged and stressed cells in a mouse model of Hutchinson-Gilford progeria syndrome (HGPS). Mice showed reduced expression of proinflammatory cytokines and extended life spans upon aged cell-specific OSK expression. The bone marrow and spleen, in particular, showed pronounced gene expression changes, and partial reprogramming in aged HGPS mice led to a shift in the cellular composition of the hematopoietic stem cell compartment toward that of young mice. Administration of AAVs carrying Cdkn2a-OSK to naturally aged wild-type mice also delayed aging phenotypes and extended life spans without altering the incidence of tumor development. Furthermore, intradermal injection of AAVs carrying Cdkn2a-OSK led to improved wound healing in aged wild-type mice. Expression of CDKN2A-OSK in aging or stressed human primary fibroblasts led to reduced expression of inflammation-related genes but did not alter the expression of cell cycle-related genes. This targeted partial reprogramming approach may therefore facilitate the development of strategies to improve health and life span and enhance resilience in the elderly.

Is the Transcription Factor NANOG Involved in Placental Aging?

PROBLEM: Accelerated placental aging is linked to abnormal fetal growth, preeclampsia (PE), and preterm birth (PTB). NANOG, a transcription factor, is known for its role in cellular reprogramming, self-renewal, and clonogenic growth. Its expression is regulated by Kruppel-like factor 4 (KLF4), which functions as both a transcriptional activator and repressor. This study evaluated the KLF4-NANOG pathway in placental samples from normal pregnancies (NP) as well as those with PE, fetal growth restriction (FGR), and PTB. METHOD OF STUDY: Placental samples from NP pregnancies and those with PE, FGR, and PTB were analyzed for NANOG and KLF4 expression using western blotting and immunohistochemistry. RESULTS: NANOG protein expression was significantly increased in placentas from PE, FGR, and PTB compared to NP (fold changes vs. NP: PE 2.48 ± 0.3, p = 0.002; FGR 1.64 ± 0.16, p = 0.03; PTB 6.03 ± 3.35, p = 0.01). Similarly, KLF4 protein expression was elevated in PE, FGR, and PTB placentas compared to NP (fold changes vs. NP: PE 5.78 ± 0.73, p = 0.001; FGR 2.61 ± 0.43, p = 0.02; PTB 11.42 ± 2.76, p = 0.0006). Immunohistochemistry revealed strong NANOG staining in the syncytiotrophoblast tissue of PE, FGR, and PTB samples, especially in extravillous trophoblasts, compared to NP placentas. CONCLUSIONS: The elevated expression of NANOG and KLF4 in abnormal placental tissues suggests their potential role as markers of enhanced placental aging and dysfunction. These findings underscore the importance of the KLF4-NANOG pathway in the pathology of PE, FGR, and PTB, providing a basis for future research into therapeutic targets for these conditions.

Endothelial γ-protocadherins inhibit KLF2 and KLF4 to promote atherosclerosis.

Atherosclerotic cardiovascular disease (ASCVD) is the leading cause of mortality worldwide. Laminar shear stress from blood flow, sensed by vascular endothelial cells, protects from ASCVD by upregulating the transcription factors KLF2 and KLF4, which induces an anti-inflammatory program that promotes vascular resilience. Here we identify clustered γ-protocadherins as therapeutically targetable, potent KLF2 and KLF4 suppressors whose upregulation contributes to ASCVD. Mechanistic studies show that γ-protocadherin cleavage results in translocation of the conserved intracellular domain to the nucleus where it physically associates with and suppresses signaling by the Notch intracellular domain. γ-Protocadherins are elevated in human ASCVD endothelium; their genetic deletion or antibody blockade protects from ASCVD in mice without detectably compromising host defense against bacterial or viral infection. These results elucidate a fundamental mechanism of vascular inflammation and reveal a method to target the endothelium rather than the immune system as a protective strategy in ASCVD.

Construction of Lentiviral Vectors Carrying Six Pluripotency Genes in Yak to Obtain Yak iPSC Cells.

Yak is an excellent germplasm resource on the Tibetan Plateau and is able to live in high-altitude areas with hypoxic, cold, and harsh environments. Studies on induced pluripotent stem cells (iPSCs) in large ruminants commonly involve a combination strategy involving six transcription factors, Oct4, Sox2, Klf4, c-Myc, Nanog, and Lin28 (OSKMNL). This strategy tends to utilize genes from the same species to optimize pluripotency maintenance. In this study, we cloned the six pluripotency genes (OSKMNL) from yak and constructed a multi-cistronic lentiviral vector carrying these genes. This vector efficiently delivered the genes into yak fibroblasts, aiming to promote the reprogramming process. We verified that the treated cells had several pluripotency characteristics, marking the first successful construction of a lentiviral system carrying yak pluripotency genes. This achievement lays the foundation for subsequent establishment of yak iPSCs and holds significant implications for yak-breed improvement and germplasm-resource conservation.

HMGA1 stimulates cancer stem-like features and sensitivity to monensin in gastric cancer.

Gastric cancer represents a serious health problem worldwide, with insufficient molecular biomarkers and therapeutic options. Consequently, several efforts have been directed towards finding specific disease markers in order to develop new therapies capable of defeating gastric cancer. Attention has been pointed to cancer stem cells (CSCs) as they are primarily responsible for tumor initiation and recurrence, making them essential therapeutic targets. Using the SORE6-GFP reporter system, based on the expression of SOX2 and/or OCT4 to drive GFP expression, we isolated gastric cancer stem-like cells (SORE6+ cells) enriched in several molecules, including SOX2, C-MYC, KLF4, HIF-1α, NOTCH1 and HMGA1. Here, we explored the previously undisclosed link of HMGA1 with gastric CSCs. Our results indicated that HMGA1 can activate a transcriptional program that includes SOX2, C-MYC, and KLF4 and endows cells with CSC features. We further showed that chemical induction of gastric CSCs using ciclopirox (CPX) can be mediated by HMGA1. Finally, we showed that HMGA1 GFP+ cells were sensitive to monensin confirming the selective activity of this drug over CSCs. Thus, HMGA1 is a key player in the cellular reprogramming of gastric non-CSCs to cancer stem-like cells.