Search Results (381)

The hypothalamus–pituitary–ovarian (HPO) axis orchestrates reproductive functions through intricate neuroendocrine crosstalk. Here, we integrated single-nucleus RNA sequencing (snRNA-seq) and spatial transcriptomics (ST) to decode the cellular heterogeneity and intercellular communication networks in the reproductive systems of pregnant Mongolian cattle. We retained a total of 6161 high-quality nuclei from the hypothalamus, 14,715 nuclei from the pituitary, and 26,072 nuclei from the ovary, providing a comprehensive cellular atlas across the HPO axis. In the hypothalamus, neurons exhibited synaptic and neuroendocrine specialization, with glutamatergic subtype Glut4 serving as a TGFβ signaling hub to regulate pituitary feedback, while GABAergic GABA1 dominated PRL signaling, likely adapting maternal behavior. Pituitary stem cells dynamically replenished endocrine populations via TGFβ, and lactotrophs formed a PRL–PRLR paracrine network with stem cells, synergizing mammary development. Ovarian luteal cells exhibited steroidogenic specialization and microenvironmental synergy: endothelial cells coregulated TGFβ-driven angiogenesis and immune tolerance, while luteal–stromal PRL–PRLR interactions amplified progesterone synthesis and nutrient support. Granulosa cells (GCs) displayed spatial-functional stratification, with steroidogenic GCs persisting across pseudotime as luteinization precursors, while atretic GCs underwent apoptosis. Spatial mapping revealed GCs’ annular follicular distribution, mediating oocyte–somatic crosstalk, and luteal–endothelial colocalization supporting vascularization. This study unveils pregnancy-specific HPO axis regulation, emphasizing multi-organ crosstalk through TGFβ/PRL pathways and stem cell-driven plasticity, offering insights into reproductive homeostasis and pathologies. Full article

Titanium–aluminum–vanadium (Ti6Al4V) is a material chosen for spine, orthopedic, and dental implants due to its combination of desirable mechanical and biological properties. Lasers have been used to modify metal surfaces, enabling the generation of a surface on Ti6Al4V with distinct micro- and nano-scale structures. Studies indicate that topography with micro/nano features of osteoclast resorption pits causes bone marrow stromal cells (MSCs) and osteoprogenitor cells to favor differentiation into an osteoblastic phenotype. This study examined whether the biological response of human MSCs to Ti6Al4V surfaces is sensitive to laser treatment-controlled micro/nano-topography. First, 15 mm diameter Ti6Al4V discs (Spine Wave Inc., Shelton, CT, USA) were either machined (M) or additively manufactured (AM). Surface treatments included no laser treatment (NT), nanosecond laser (Ns), femtosecond laser (Fs), or nanosecond followed by femtosecond laser (Ns+Fs). Surface wettability, roughness, and surface chemistry were determined using sessile drop contact angle, laser confocal microscopy, X-ray photoelectron spectroscopy (XPS), and scanning electron microscopy (SEM). Human MSCs were cultured in growth media on tissue culture polystyrene (TCPS) or test surfaces. On day 7, the levels of osteocalcin (OCN), osteopontin (OPN), osteoprotegerin (OPG), and vascular endothelial growth factor 165 (VEGF) in the conditioned media were measured. M NT, Fs, and Ns+Fs surfaces were hydrophilic; Ns was hydrophobic. AM NT and Fs surfaces were hydrophilic; AM Ns and Ns+Fs were hydrophobic. Roughness (Sa and Sz) increased after Ns and Ns+Fs treatment for both M and AM disks. All surfaces primarily consisted of oxygen, titanium, and carbon; Fs had increased levels of aluminum for both M and AM. SEM images showed that M NT discs had a smooth surface, whereas AM surfaces appeared rough at a higher magnification. Fs surfaces had a similar morphology to their respective NT disc at low magnification, but higher magnification revealed nano-scale bumps not seen on NT surfaces. AM Fs surfaces also had regular interval ridges that were not seen on non-femto laser-ablated surfaces. Surface roughness was increased on M and AM Ns and Ns+Fs disks compared to NT and Fs disks. OCN was enhanced, and DNA was reduced on Ns and Ns+Fs, with no difference between them. OPN, OPG, and VEGF levels for laser-treated M surfaces were unchanged compared to NT, apart from an increase in OPG on Fs. MSCs grown on AM Ns and Ns+Fs surfaces had increased levels of OCN per DNA. These results indicate that MSCs cultured on AM Ns and AM Ns+Fs surfaces, which exhibited unique roughness at the microscale and nanoscale, had enhanced differentiation to an osteoblastic phenotype. The laser treatments of the surface mediated this enhancement of MSC differentiation and warrant further clinical investigation. Full article

Cardiac tissue engineering is a promising strategy to restore cardiac function in heart failure patients. Understanding the cardiac tissue architecture including the cardiac stroma is essential for developing not only advanced cardiac tissue engineering but also novel therapeutic strategies. One of the crucial components of the cardiac stroma is the myocardial vasculature. To enhance the spatial visualization of the cardiac stromal cytoarchitecture with a particular focus on myocardial vasculature, we performed 3D reconstructions of the murine cardiac micro vessels using Serial Block-Face Scanning Electron Microscopy (SBF-SEM). These analyses revealed that pericyte cell bodies were primarily oriented lengthwise and extended several cellular protrusions towards the endothelium. At capillary branching points, some pericytes made contact with both capillaries emerging from branching. In addition to pericytes that are completely encapsulated by the common basal lamina together with capillary endothelial cells, we identified other vascular-associated cells located outside this sheath. Based on marker expression, these cells were distinguished from fibroblasts and suggested to be telocytes. The vascular-associated cells formed electron-dense contact zones with endothelial cells, suggesting functional coupling between these both cell types. In conclusion, this study provides detailed three-dimensional visualizations of the cardiac stroma with a particular focus on cardiac microvasculature, offering enhanced insight into the cardiac stromal cytoarchitecture. Full article

Micro-fragmented adipose tissue (MFAT) is a promising autologous therapy for knee osteoarthritis. To avoid repeated liposuction procedures for its clinical application, MFAT obtained from patients with knee osteoarthritis was stored at −80 °C in a tissue bank. This study describes the preparation, cryopreservation, thawing, and washing, as well as comprehensive analysis of cell populations in fresh and MFAT thawed after two years. Immunophenotyping of both fresh and thawed MFAT showed a significant presence of endothelial progenitors and pericytes in the stromal vascular fraction. Viability before (59.75%) and after freezing (55.73%) showed no significant difference. However, the average cell count per gram of MFAT was significantly reduced in thawed samples (3.00 × 10⁵) compared to fresh ones (5.64 × 10⁵), likely due to processing steps. Thawed MFAT samples showed increased CD73 expression on the CD31^highCD34^high subset of EP and SA-ASC, as well as increased expression of CD105 on EP, the CD31^lowCD34^low subset of EP, pericytes, and SA-ASC. Microbiological testing confirmed 100% sterility, and double washing efficiently removed DMSO, confirming sample safety. Histological analysis revealed healthy, uniformly shaped adipocytes with intact membranes. This approach allows accurate estimation of cell yield for intra-articular injection, ensuring delivery of the target cell number into the knee. Quality control analysis confirms that cryopreserved MFAT retains high cellular and structural integrity, supporting its safety and suitability for clinical application. Full article

Current treatment strategies for partial tendon tears often lack the capacity to promote true tissue regeneration and improve long-term clinical outcomes. This study tested the hypothesis that treatment of a partial defect in the rabbit common calcaneus tendon (CCT) with uncultured, unmodified, autologous, adipose-derived regenerative cells (UA-ADRCs) enables regenerative healing without scar formation. A full-thickness, 3 mm defect was produced in the midsubstance of the right gastrocnemius tendon, a component of the CCT, in adult female New Zealand white rabbits. Animals received either an injection of 28.3 × 10⁶ UA-ADRCs in 0.5 mL Ringer’s lactated solution (RLS) or saline, or RLS or saline alone as sham treatment. Tendons were analyzed 4 or 12 weeks post-treatment using histology, immunohistochemistry and non-destructive biomechanical testing. UA-ADRC-treated tendons showed newly formed connective tissue consistent with tendon regeneration, whereas sham-treated tendons developed scar tissue. Biomechanical testing showed significantly higher percent relaxation in UA-ADRC-treated tendons compared to sham controls (p < 0.05), indicating greater viscoelasticity characteristic of healthy or well-integrated tissue. Together, these findings suggest that UA-ADRC therapy may provide a regenerative, structure-modifying treatment for partial tendon tears. Full article

Tissue-engineered artificial skin has the potential to enhance wound healing without necessitating extensive surgical procedures or causing donor-site morbidity. The purpose of this study was to examine the possibility of developing tri-layered tissue-engineered full-thickness artificial skin with a basement membrane for clinical use to accelerate wound healing. We engineered full-thickness artificial skin with a basement membrane for wound healing by employing stromal vascular fraction (SVF) cells for the dermal layer and autologous keratinocytes for the epidermal layer. The fabrication of a basement membrane involved the use of 100% bovine collagen and 4% elastin produced through a low-temperature three-dimensional printer. Scaffolds for cells were printed with 100% bovine collagen. The basement membrane underwent evaluations for collagenase degradation, tensile strength, and structural characteristics using scanning electron microscopy. The final tri-layered full-thickness artificial skin included two cell scaffolds with a basement membrane between them. The basement membrane may support cellular attachment without inducing significant cytotoxic effects. This study presents a novel strategy for full-thickness artificial skin development, combining SVF and keratinocytes with an optimized collagen-elastin basement membrane. This method may overcome the significant limitations of current artificial skin, thereby contributing to the advancement of tissue-engineering in wound healing for clinical use. Full article

We developed biomimetic full-thickness artificial skin using stromal vascular fraction (SVF) cells and autologous keratinocytes for the dermal and epidermal layers of skin, respectively. Full-thickness artificial skin scaffolds were fabricated using 4% porcine collagen and/or elastin in a low-temperature three-dimensional printer. Two types of scaffolds with collagen-to-elastin ratios of 100:0 and 100:4 were printed and compared. The scaffolds were analyzed for collagenase degradation, tensile strength, and structural features using scanning electron microscopy. By 24 h, the collagen-only scaffolds showed gradual degradation, and the collagen-elastin scaffolds retained the highest structural integrity but were not degraded. In the tensile strength tests, the collagen-only scaffolds exhibited a tensile strength of 2.2 N, while the collagen-elastin scaffolds showed a tensile strength of 4.2 N. Cell viability tests for keratinocytes displayed an initial viability of 89.32 ± 3.01% on day 1, which gradually increased to 97.22 ± 4.99% by day 7. Similarly, SVF cells exhibited a viability of 93.68 ± 1.82% on day 1, which slightly improved to 97.12 ± 1.64% on day 7. This study presents a novel strategy for full-thickness artificial skin development, combining SVF and keratinocytes with an optimized single collagen scaffold and a gradient pore-density structure. Full article

T-cadherin (CDH13) is an atypical, glycosyl-phosphatidylinositol-anchored cadherin with functions ranging from axon guidance and vascular patterning to adipokine signaling and cell-fate specification. Originally identified as a homophilic cue for migrating neural crest cells, projecting axons, and growing blood vessels, it later emerged as a dual metabolic receptor for cardioprotective high-molecular-weight adiponectin and atherogenic low-density lipoproteins. We recently showed that mesenchymal stem/stromal cells lacking T-cadherin are predisposed to adipogenesis, underscoring its role in lineage choice. Emerging evidence indicates that CDH13 expression and function are fine-tuned by non-coding RNAs (ncRNAs). MiR-199b-5p, miR-377-3p, miR-23a/27a/24-2, and the miR-142 family directly bind CDH13 3′-UTR or its epigenetic regulators, affecting transcription or accelerating decay. Long non-coding RNAs (lncRNAs), including antisense transcripts CDH13-AS1/AS2, brain-restricted FEDORA, and context-dependent LINC00707 and UPAT, either sponge these miRNAs or recruit DNMT/TET enzymes to the CDH13 promoter. Circular RNAs (circRNAs), i.e.circCDH13 and circ_0000119, can add a third level of complexity by sequestering miRNA repressors or boosting DNMT1. Collectively, this ncRNA circuitry regulates T-cadherin across cardiovascular, metabolic, oncogenic, and neurodegenerative conditions. This review integrates both experimentally validated data and in silico predictions to map the ncRNA-CDH13 crosstalk between health and disease, opening new avenues for biomarker discovery and RNA-based therapeutics. Full article

Plantar fasciitis is a common condition characterized by inflammation and degeneration of the plantar fascia, leading to heel pain and reduced mobility. Affecting both athletic and non-athletic populations, it is a leading cause of foot-related medical visits. Conservative treatments, including rest, physical therapy, and corticosteroid injections, provide relief for most patients, but a subset experiences persistent symptoms requiring advanced therapies. Emerging biologic treatments, such as platelet-rich plasma (PRP) and mesenchymal stem/stromal cell (MSC) therapy, have demonstrated potential in promoting tissue regeneration and reducing inflammation. Recently, MSC-derived extracellular vesicles (MSC-EVs) have gained attention for their regenerative properties, offering a promising, cell-free therapeutic approach. EVs mediate tissue repair through immunomodulation, anti-inflammatory signaling, and extracellular matrix stabilization. Preclinical studies suggest that EV therapy may improve tendon and ligament healing by promoting M2 macrophage polarization, inhibiting excessive metalloproteinase activity, and enhancing vascular remodeling. This review explores the potential of MSC-EVs as an innovative, non-surgical treatment for plantar fasciitis, addressing their mechanisms of action and current evidence in musculoskeletal regeneration. Full article

Cartilage and bone defects as well as osteoarthritis are prevalent worldwide, affecting individuals across all age groups, from young, active populations to older adults. The standard protocol in cartilage regeneration involves knee replacement surgery through the implantation of an endoprosthesis. Current clinical protocols involving cell-based therapies are associated with limitations, including the lack of functional cartilage-like tissue and dedifferentiation of chondrocyte, particularly during monoculture. Similarly, in bone regeneration, the “gold standard” is the use of bone auto- or allografts, which are associated with immunological rejection, inadequate vascularization, and limited osteogenesis. To overcome these limitations, various co-culture techniques have been introduced as promising strategies for cartilage and bone tissue regeneration. These systems aim to mimic native microenvironments by promoting interactions between chondrocytes and mesenchymal stromal cells (MSCs) in cartilage repair and between osteogenic and angiogenic cells in bone regeneration. This paper introduces different co-culture systems focusing on in vitro crosstalk between MSCs derived from various sources and other somatic cell populations in cartilage and bone regeneration. Full article

Background: Histological and immunohistochemical analyses of adipose tissue are essential for evaluating the quality and functionality of lipoaspirates in regenerative medicine and fat grafting procedures. These methods provide insights into tissue viability, cellular subtypes, and extracellular matrix (ECM) composition—all factors influencing graft retention and clinical outcomes. Purpose: This scoping review aims to summarize the most commonly used staining methods and their applications in the histology and immunohistochemistry of adipose tissue. By exploring qualitative and quantitative markers, we seek to guide researchers in selecting the appropriate methodologies for addressing experimental and translational research. Methods: A systematic search was conducted using PubMed, Ovid, and the Cochrane Library databases from inception to 2024, employing Boolean operators (“lipoaspirate” OR “fat graft” OR “gauze rolling” OR “decantation” OR “coleman fat” OR “celt” OR “nanofat” OR “lipofilling” OR “human fat” AND “histol*”). Studies were included if they utilized histology or immunohistochemistry on undigested human adipose tissue or its derivatives. The inclusion criteria focused on peer-reviewed, English-language studies reporting quantitative and qualitative data on adipose tissue markers. Results: Out of 166 studies analyzed, hematoxylin–eosin (H&E) was the most frequently employed histological stain (152 studies), followed by Masson Trichrome and Sudan III. Immunohistochemical markers such as CD31, CD34, and perilipin were extensively used to distinguish stromal vascular fraction (SVF) cells, adipocytes, and inflammatory processes. Studies employing semiquantitative scoring demonstrated enhanced comparability, particularly for fibrosis, necrosis, and oil cyst evaluation. Quantitative analyses focused on SVF cell density, mature adipocyte integrity, and ECM composition. Methodological inconsistencies, particularly in preparation protocols, were observed in 25 studies. Conclusions: This review highlights the critical role of histological and immunohistochemical methods in adipose tissue research. H&E staining remains the cornerstone for general tissue evaluation in the clinical context, while specialized stains and immunohistochemical markers allow for detailed analyses of specific cellular and ECM components in experimental research. Standardizing preparation and evaluation protocols will enhance interstudy comparability and support advancements in adipose tissue-based therapies. Full article

Endometrial organoids (EOs) have emerged as a powerful three-dimensional (3D) model for studying the human endometrium, offering new insights into infertility and reproductive disorders. These self-organizing miniature structures closely mimic the cellular composition, hormonal responsiveness, and functional characteristics of the endometrium, making them valuable preclinical tools for investigating implantation failure, endometrial receptivity, and disease pathophysiology. This review explores the role of EOs in reproductive medicine, with a focus on their applications in infertility research, environmental toxicology, and regenerative therapies. Traditional 2D cell cultures fail to capture the complexity of these physiological and pathological interactions, whereas organoids provide a physiologically relevant system for studying implantation mechanisms. Additionally, co-culture models incorporating stromal and immune cells have further enhanced our understanding of the maternal–fetal interface. Beyond modeling infertility, EOs hold significant promise for therapeutic applications. Advances in organoid transplantation have demonstrated potential for treating endometrial dysfunction-related infertility, including conditions such as Asherman’s syndrome and thin endometrium. Moreover, these models serve as a platform for drug screening and biomarker discovery, paving the way for personalized reproductive medicine. Despite their transformative potential, limitations remain, including the need for improved extracellular matrices, vascularization, and immune system integration. This review emphasizes the significant contributions of EOs to the field of infertility treatment and reproductive biology by examining recent advancements and emerging research. The continued refinement of these models would offer a paradigm for improving assisted reproductive technologies (ARTs) and regenerative medicine outcomes, offering new hope for individuals facing infertility challenges. Full article

Aging disrupts the bone marrow (BM) niche, a complex microenvironment crucial for hematopoietic stem cell (HSC) maintenance. A key, yet debated, hallmark of this aging process is the accumulation of bone marrow adipocytes (BMAds). This review explores the evolving role of BMAds in the aging BM, particularly their influence on HSC regulation via metabolic, endocrine, and inflammatory pathways. Aging BMAds exhibit altered secretory profiles, including reduced leptin and adiponectin and increased pro-inflammatory signals, which skew hematopoiesis toward myeloid over lymphoid lineage production. Additionally, shifts in fatty acid composition and lactate signaling from BMAds may impair stem cell function. These changes, alongside aging-associated alterations in vascular, neural, and stromal components of the niche, contribute to diminished immune resilience in older adults. We discuss emerging therapeutic strategies targeting BMAd-derived factors, such as DPP4 inhibition or the modulation of β-adrenergic signaling, aimed at creating a more youthful BM environment. By summarizing current insights into the aging BM niche and the central role of BMAds, this review highlights mechanisms that could be targeted to rejuvenate hematopoiesis and improve immune function in the elderly. Full article

Papillary thyroid cancer (PTC) contains mesenchymal stem/stromal cells (MSCs), but their contribution to PTC progression is not clear. In this study, we compared the transcriptional signatures of normal thyroid (NT) and PTC-derived MSCs with the aim of determining if these have distinct transcriptomes that might influence PTC progression. We used flow cytometry in combination with a panel of MSC clusters of differentiation (CD) markers and showed that both thyroid MSC populations expressed MSC markers and lacked expression of markers not normally expressed by MSCs. In addition, we determined that both MSC populations could differentiate to adipocytes and osteocytes. Analysis of single cell RNA sequencing data from both MSC populations revealed, regardless of tissue of origin, that both contained similar numbers of subpopulations. Cluster analysis revealed similarity in expression of both MSC populations for stromal markers, the vascular marker VEGFA and the smooth muscle marker CALD1, while smaller subpopulations expressed markers of more lineage-committed thyroid cells. PTC MSCs also showed upregulated expression of 28 genes, many of which are known to be involved in epithelial–mesenchymal transition (EMT) and/or disease progression in several types of cancers, including but not limited to breast cancer, gastric cancer, cervical carcinoma, bladder cancer and thyroid cancer. This included several members of the S100 and IGFBP gene families. Taken together, these data support a role for PTC MSCs in PTC progression. Full article

Background: Von Hippel-Lindau (VHL) disease, a hereditary cancer syndrome, is characterized by mutations in the VHL gene, which result in the stabilization of hypoxia-inducible factors (HIF)-1α and -2α, ultimately leading to the development of highly vascularized tumors, such as hemangioblastomas of the central nervous system (CNS-HBs). The standard treatment for these brain tumors is neurosurgical resection. However, multiple surgeries are often necessary due to tumor recurrence, which increases the risk of neurological sequelae. Thus, elucidation of the proliferative behavior of hemangioblastomas (with the aim of identifying biomarkers associated with tumor progression) and the development of pharmacological therapies could reduce the need for repeated surgical interventions and provide alternative treatment options for unresectable CNS-HBs. Belzutifan (Welireg™), a selective HIF-2α inhibitor and the only FDA-approved non-surgical option, has shown limited efficacy in CNS-HBs, highlighting the need for alternative therapeutic strategies. Results: In this study, primary cell cultures were successfully established from CNS-HB tissue samples of VHL patients, achieving a 75% success rate. These cultures were predominantly composed of stromal cells and pericytes. The proliferative patterns of patient-derived HB cell cultures significantly correlated with tumor burden and recurrence in VHL patients. Furthermore, flow cytometry, reverse transcription-PCR, and Western blot analyses revealed marked overexpression of both HIF-1α and HIF-2α isoforms in primary HB cells. In addition, evaluation of the therapeutic potential of acriflavine, a dual HIF-1α/HIF-2α inhibitor, demonstrated reduced HB cells viability, induced G2/M cell cycle arrest, and predominantly triggered necrotic cell death in patient-derived HB cultures. Conclusions: These results suggest that the in vitro proliferative dynamics of HB cell cultures may reflect clinical characteristics associated with CNS-HB progression, potentially serving as indicators to predict tumor development in patients with VHL. Furthermore, our findings support the simultaneous targeting of both HIF-1α and HIF-2α isoforms as a promising non-invasive therapeutic strategy. Full article