Molecular Morphology and Function of Stromal Cells.

The term "stromal cells" refers to a highly heterogeneous class of connective tissue cells that build the infrastructure of any organ and fulfill a variety of fundamental roles in health and disease [...].

Fibroblast dedifferentiation as a determinant of successful regeneration.

Limb regeneration, while observed lifelong in salamanders, is restricted in post-metamorphic Xenopus laevis frogs. Whether this loss is due to systemic factors or an intrinsic incapability of cells to form competent stem cells has been unclear. Here, we use genetic fate mapping to establish that connective tissue (CT) cells form the post-metamorphic frog blastema, as in the case of axolotls. Using heterochronic transplantation into the limb bud and single-cell transcriptomic profiling, we show that axolotl CT cells dedifferentiate and integrate to form lineages, including cartilage. In contrast, frog blastema CT cells do not fully re-express the limb bud progenitor program, even when transplanted into the limb bud. Correspondingly, transplanted cells contribute to extraskeletal CT, but not to the developing cartilage. Furthermore, using single-cell RNA-seq analysis we find that embryonic and adult frog cartilage differentiation programs are molecularly distinct. This work defines intrinsic restrictions in CT dedifferentiation as a limitation in adult regeneration.

Patient Age and Cell Concentration Influence Prevalence and Concentration of Progenitors in Bone Marrow Aspirates: An Analysis of 436 Patients.

BACKGROUND: Connective tissue progenitors (CTPs) resident in native tissues serve as biological building blocks in tissue repair and remodeling processes. Methods for analysis and reporting on CTP quantity and quality are essential for defining optimal cell sources and donor characteristics and the impact of cell processing methods for cell therapy applications. The present study examines the influence of donor characteristics and cell concentration (nucleated cells/mL) on CTP prevalence (CTPs/million nucleated cells) and CTP concentration (CTPs/mL) in bone marrow aspirates (BMAs). METHODS: Iliac crest bone marrow was aspirated from 436 patients during elective total knee or hip arthroplasty. Bone marrow-derived nucleated cells were plated at a density of 1.19 × 105 cells/cm2. Colony-forming unit analysis was performed on day 6. RESULTS: Large variation was seen between donors. Age (p < 0.05) and cell concentration (p < 0.001) significantly influenced CTP prevalence and CTP concentration. For every 1-year increase in age, the odds of having at least an average CTP prevalence and CTP concentration decreased by 1.5% and 1.6%, respectively. For every 1 million cells/mL increase in cell concentration, the odds of having at least an average CTP prevalence and CTP concentration increased by 2.2% and 7.9%, respectively. Sex, race, body mass index (BMI), and the presence of osteoporosis did not influence CTP prevalence or CTP concentration. CONCLUSIONS: BMA-derived CTPs were obtained from all patient groups. CTP prevalence and CTP concentration decreased with age. Cell concentration decreased with age and positively correlated with total CTP prevalence and CTP concentration. The mean CTP concentration in patients >60 years of age was a third of the CTP concentration in patients <30 years of age. CLINICAL RELEVANCE: Proper BMA techniques are necessary to obtain a high-quality yield and composition of cells and CTPs. The reduced CTP concentration and CTP prevalence in the elderly may be mitigated by the use of cell processing methods that increase CTP concentration and CTP prevalence (e.g., by removing red blood cells, serum, and non-CTPs or by increasing aspirate volumes). Cell concentration in the BMA can be measured at the point of care and is an appropriate initial assessment of the quality of BMA.

MUSE Stem Cells Can Be Isolated from Stromal Compartment of Mouse Bone Marrow, Adipose Tissue, and Ear Connective Tissue: A Comparative Study of Their In Vitro Properties.

The cells present in the stromal compartment of many tissues are a heterogeneous population containing stem cells, progenitor cells, fibroblasts, and other stromal cells. A SSEA3(+) cell subpopulation isolated from human stromal compartments showed stem cell properties. These cells, known as multilineage-differentiating stress-enduring (MUSE) cells, are capable of resisting stress and possess an excellent ability to repair DNA damage. We isolated MUSE cells from different mouse stromal compartments, such as those present in bone marrow, subcutaneous white adipose tissue, and ear connective tissue. These cells showed overlapping in vitro biological properties. The mouse MUSE cells were positive for stemness markers such as SOX2, OCT3/4, and NANOG. They also expressed TERT, the catalytic telomerase subunit. The mouse MUSE cells showed spontaneous commitment to differentiation in meso/ecto/endodermal derivatives. The demonstration that multilineage stem cells can be isolated from an animal model, such as the mouse, could offer a valid alternative to the use of other stem cells for disease studies and envisage of cellular therapies.

New inducible mast cell-deficient mouse model (Mcpt5/Cma1DTR).

Mast cell-deficient mice are helpful for understanding the roles of mast cells in vivo. To date, a dozen mouse models for mast cell deficiency have been reported. However, mice with a specific depletion of all populations of mast cells have not been reported. We generated knock-in mice, termed Mcpt5/Cma1DTR mice, expressing human diphtheria toxin A (DT) receptor under the endogenous promoter of Mcpt5 (also known as Cma1), which encodes mouse mast cell protease-5. Flow cytometry and histological analysis showed that intraperitoneal injection of DT induced almost complete depletion of mast cells in heterozygote Mcpt5/Cma1DTR/+ mice. The deletion rates of mast cells in peritoneal cavity, mesentery, abdominal skin, ear skin, and glandular stomach were 99.9%, 100%, 98.7%, 97.7%, and 100%, respectively. Passive cutaneous anaphylaxis reaction also revealed mast cell deficiency in ear skin after DT treatment. Other than mast cells, a small percentage of marginal zone B cells in Mcpt5/Cma1DTR/+ mice were killed by DT treatment. In conclusion, the Mcpt5/Cma1DTR/+ mouse model is valuable for achieving conditional depletion of all populations of mast cells without inducing a marked reduction in other cells.

Characterization of connective tissues using near-infrared spectroscopy and imaging.

Near-infrared (NIR) spectroscopy is a powerful analytical method for rapid, non-destructive and label-free assessment of biological materials. Compared to mid-infrared spectroscopy, NIR spectroscopy excels in penetration depth, allowing intact biological tissue assessment, albeit at the cost of reduced molecular specificity. Furthermore, it is relatively safe compared to Raman spectroscopy, with no risk of laser-induced photothermal damage. A typical NIR spectroscopy workflow for biological tissue characterization involves sample preparation, spectral acquisition, pre-processing and analysis. The resulting spectrum embeds intrinsic information on the tissue's biomolecular, structural and functional properties. Here we demonstrate the analytical power of NIR spectroscopy for exploratory and diagnostic applications by providing instructions for acquiring NIR spectra, maps and images in biological tissues. By adapting and extending this protocol from the demonstrated application in connective tissues to other biological tissues, we expect that a typical NIR spectroscopic study can be performed by a non-specialist user to characterize biological tissues in basic research or clinical settings. We also describe how to use this protocol for exploratory study on connective tissues, including differentiating among ligament types, non-destructively monitoring changes in matrix formation during engineered cartilage development, mapping articular cartilage proteoglycan content across bovine patella and spectral imaging across the depth-wise zones of articular cartilage and subchondral bone. Depending on acquisition mode and experiment objectives, a typical exploratory study can be completed within 6 h, including sample preparation and data analysis.

Diversity in cancer invasion phenotypes indicates specific stroma regulated programs.

Tumor dissemination into the surrounding stroma is the initial step in a metastatic cascade. Invasion into stroma is a non-autonomous process for cancer, and its progression depends upon the stage of cancer, as well as the cells residing in the stroma. However, a systems framework to understand how stromal fibroblasts resist, collude, or aid cancer invasion has been lacking, limiting our understanding of the role of stromal biology in cancer metastasis. We and others have shown that gene perturbation in stromal fibroblasts can modulate cancer invasion into the stroma, highlighting the active role stroma plays in regulating its own invasion. However, cancer invasion into stroma is a complex higher-order process and consists of various sub-phenotypes that together can result in an invasion. Stromal invasion exhibits a diversity of modalities in vivo. It is not well understood if these diverse modalities are correlated, or they emanate from distinct mechanisms and if stromal biology could regulate these characteristics. These characteristics include the extent of invasion, formation, and persistence of invasive forks by cancer as opposed to a collective frontal invasion, the persistence of invading velocity by leader cells at the tip of invasive forks, etc. We posit that quantifying distinct aspects of collective invasion can provide useful suggestions about the plausible mechanisms regulating these processes, including whether the process is regulated by mechanics or by intercellular communication between stromal cells and cancer. Here, we have identified the sub-characteristics of invasion, which might be indicative of broader mechanisms regulating these processes, developed methods to quantify these metrics, and demonstrated that perturbation of stromal genes can modulate distinct aspects of collective invasion. Our results highlight that the genetic state of stromal fibroblasts can regulate complex phenomena involved in cancer dissemination and suggest that collective cancer invasion into stroma is an outcome of the complex interplay between cancer and stromal fibroblasts.

Individual Limb Muscle Bundles Are Formed through Progressive Steps Orchestrated by Adjacent Connective Tissue Cells during Primary Myogenesis.

Although the factors regulating muscle cell differentiation are well described, we know very little about how differentiating muscle fibers are organized into individual muscle tissue bundles. Disruption of these processes leads to muscle hypoplasia or dysplasia, and replicating these events is vital in tissue engineering approaches. We describe the progressive cellular events that orchestrate the formation of individual limb muscle bundles and directly demonstrate the role of the connective tissue cells that surround muscle precursors in controlling these events. We show how disruption of gene activity within or genetic ablation of connective tissue cells impacts muscle precursors causing disruption of muscle bundle formation and subsequent muscle dysplasia and hypoplasia. We identify several markers of the populations of connective tissue cells that surround muscle precursors and provide a model for how matrix-modifying proteoglycans secreted by these cells may influence muscle bundle formation by effects on the local extracellular matrix (ECM) environment.

A journal of histochemistry as a forum for non-histochemical scientific societies.

Histochemical techniques are widely applied in biomedical research and, during the last twenty years, they were among the methods used in more than 590,000 scientific articles in indexed journals. However, a very small percentage of these papers were published in strictly histochemical journals. A possible strategy to widen the audience of the histochemical journals making them attractive to non-histochemist authors might be to publish and make open-access available the proceedings of the meetings and conferences of valued scientific societies whose fellows use microscopy and histochemistry in their experimental activity. In the last years' experience of the European Journal of Histochemistry, this approach was effective to increase the number of published articles on stem cells and development, connective tissue and nerve cell biology.

Autophagy in nutrient storage cells of the Pacific oyster, Crassostrea gigas.

In oysters, nutrients are stored in a special type of cells referred to as vesicular-connective tissue cells (VCT-cells). These cells accumulate and provide nutrient to satisfy various needs of the organism, including gametogenesis. During the annual reproductive cycle, VCT-cells pass through a series of changes in their morphology associated with nutrients mobilization for developing germ cells. The results presented here show an approximately 33-35% increase in the number of autophagic vesicles in cytoplasm of VCT-cells in the gonadal area of C. gigas during the stage of active gametogenesis as compared to the resting stage of reproductive cycle. No destruction of VCT-cells due to autophagy or any other factors was observed, both in males and females. Our results indicate that autophagy does increase in VCT-cells of C. gigas and plays a certain role in nutrient mobilization from these cells.

Dynamic Compressive Loading Improves Cartilage Repair in an In Vitro Model of Microfracture: Comparison of 2 Mechanical Loading Regimens on Simulated Microfracture Based on Fibrin Gel Scaffolds Encapsulating Connective Tissue Progenitor Cells.

BACKGROUND: Microfracture of focal chondral defects often produces fibrocartilage, which inconsistently integrates with the surrounding native tissue and possesses inferior mechanical properties compared with hyaline cartilage. Mechanical loading modulates cartilage during development, but it remains unclear how loads produced in the course of postoperative rehabilitation affect the formation of the new fibrocartilaginous tissue. PURPOSE: To assess the influence of different mechanical loading regimens, including dynamic compressive stress or rotational shear stress, on an in vitro model of microfracture repair based on fibrin gel scaffolds encapsulating connective tissue progenitor cells. STUDY DESIGN: Controlled laboratory study. METHODS: Cylindrical cores were made in bovine hyaline cartilage explants and filled with either (1) cartilage plug returned to original location (positive control), (2) fibrin gel (negative control), or (3) fibrin gel with encapsulated connective tissue progenitor cells (microfracture mimic). Constructs were then subjected to 1 of 3 loading regimens: (1) no loading (ie, unloaded), (2) dynamic compressive loading, or (3) rotational shear loading. On days 0, 7, 14, and 21, the integration strength between the outer chondral ring and the central insert was measured with an electroforce mechanical tester. The central core component, mimicking microfracture neotissue, was also analyzed for gene expression by real-time reverse-transcription polymerase chain reaction, glycosaminoglycan, and double-stranded DNA contents, and tissue morphology was analyzed histologically. RESULTS: Integration strengths between the outer chondral ring and central neotissue of the cartilage plug and fibrin + cells groups significantly increased upon exposure to compressive loading compared with day 0 controls (P = .007). Compressive loading upregulated expression of chondrogenesis-associated genes (SRY-related HGMG box-containing gene 9 [SOX9], collagen type II α1 [COL2A1], and increased ratio of COL2A1 to collagen type I α1 [COL1A1], an indicator of more hyaline phenotype) in the neotissue of the fibrin + cells group compared with the unloaded group at day 21 (SOX9, P = .0032; COL2A1, P < .0001; COL2A1:COL1A1, P = .0308). Fibrin + cells constructs exposed to shear loading expressed higher levels of chondrogenic genes compared with the unloaded condition, but the levels were not as high as those for the compressive loading condition. Furthermore, catabolic markers (MMP3 and ADAMTS 5) were significantly upregulated by shear loading (P = .0234 and P < .0001, respectively) at day 21 compared with day 0. CONCLUSION: Dynamic compressive loading enhanced neotissue chondrogenesis and maturation in a simulated in vitro model of microfracture, with generation of more hyaline-like cartilage and improved integration with the surrounding tissue. CLINICAL RELEVANCE: Controlled loading after microfracture may be beneficial in promoting the formation of more hyaline-like cartilage repair tissue; however, the loading regimens applied in this in vitro model do not yet fully reproduce the complex loading patterns created during clinical rehabilitation. Further optimization of in vitro models of cartilage repair may ultimately inform rehabilitation protocols.

Suppression of IgE-Independent Degranulation of Murine Connective Tissue-Type Mast Cells by Dexamethasone.

Steroidal anti-inflammatory drugs are widely used for the treatment of chronic cutaneous inflammation, such as atopic dermatitis, although it remains unknown how they modulate cutaneous mast cell functions. We investigated the effects of prolonged treatment with a synthetic glucocorticoid, dexamethasone, on murine connective tissue-type mast cells using in vitro and in vivo models. Our connective tissue-type bone marrow-derived cultured mast cell model was found to be sensitive to mast cell secretagogues, such as compound 48/80 and substance P, and higher expression levels of α subunit of a trimeric G protein, Gi1, and several Mas-related G protein-coupled receptor (Mrgpr) subtypes were observed in comparison with immature cultured mast cells. Secretagogue-induced degranulation and up-regulation of these genes was suppressed when cultured in the presence of dexamethasone. The profiles of granule constituents were drastically altered by dexamethasone. Topical application of dexamethasone down-modulated secretagogue-induced degranulation and the expression levels of several Mrgpr subtypes in cutaneous tissue. These results suggest that mast cell-mediated IgE-independent cutaneous inflammation could be suppressed by steroidal anti-inflammatory drugs through the down-regulation of G αi1 and several Mrgpr subtypes in mast cells.

Hyaluronic acid, CD44 and RHAMM regulate myoblast behavior during embryogenesis.

Hyaluronic acid (HA) is an extracellular matrix (ECM) component that has been shown to play a significant role in regulating muscle cell behavior during repair and regeneration. For instance, ECM remodeling after muscle injury involves an upregulation in HA expression that is coupled with skeletal muscle precursor cell recruitment. However, little is known about the role of HA during skeletal muscle development. To gain insight into the way in which HA mediates embryonic myogenesis, we first determined the spatial distribution and gene expression of CD44, RHAMM and other HA related proteins in embryonic day (E)10.5 to E12.5 murine forelimbs. While HA and CD44 expression remained high, RHAMM decreased at both the protein (via immunohistochemistry) and RNA (via qPCR) levels. Next, we determined that 4-methylumbelliferone-mediated knockdown of HA synthesis inhibited the migration and proliferation of E11.5/E12.5 forelimb-derived cells. Then, the influence of CD44 and RHAMM on myoblast and connective tissue cell behavior was investigated using antibodies against these receptors. Anti-RHAMM, but not anti-CD44, significantly decreased the total distance myogenic progenitors migrated over 24 h, whereas both inhibited connective tissue cell migration. In contrast, anti-CD44 inhibited the proliferation of connective tissue cells and muscle progenitors, but anti-RHAMM had no effect. However, when myoblasts and connective tissue cells were depleted of CD44 and RHAMM by shRNA, motility and proliferation were significantly inhibited in both cells indicating that blocking cell surface-localized CD44 and RHAMM does not have as pronounced effect as global shRNA-mediated depletion of these receptors. These results show, for the first time, the distribution and activity of RHAMM in the context of skeletal muscle. Furthermore, our data indicate that HA, through interactions with CD44 and RHAMM, promotes myogenic progenitor migration and proliferation. Confirmation of the role of HA and its receptors in directing myogenesis will be useful for the design of regenerative therapies that aim to promote the restoration of damaged or diseased muscle.

In vitro placenta barrier model using primary human trophoblasts, underlying connective tissue and vascular endothelium.

Fetal development may be compromised by adverse events at the placental interface between mother and fetus. However, it is still unclear how the communication between mother and fetus occurs through the placenta. In vitro - models of the human placental barrier, which could help our understanding and which recreate three-dimensional (3D) structures with biological functionalities and vasculatures, have not been reported yet. Here we present a 3D-vascularized human primary placental barrier model which can be constructed in 1 day. We illustrate the similarity of our model to first trimester human placenta, both in its structure and in its ability to respond to altered oxygen and to secrete factors that cause damage cells across the barrier including embryonic cortical neurons. We use this model to highlight the possibility that both the trophoblast and the endothelium within the placenta might play a role in the fetomaternal dialogue.

Variation in primary and culture-expanded cells derived from connective tissue progenitors in human bone marrow space, bone trabecular surface and adipose tissue.

BACKGROUND AIMS: Connective tissue progenitors (CTPs) embody the heterogeneous stem and progenitor cell populations present in native tissue. CTPs are essential to the formation and remodeling of connective tissue and represent key targets for tissue-engineering and cell-based therapies. To better understand and characterize CTPs, we aimed to compare the (i) concentration and prevalence, (ii) early in vitro biological behavior and (iii) expression of surface-markers and transcription factors among cells derived from marrow space (MS), trabecular surface (TS), and adipose tissues (AT). METHODS: Cancellous-bone and subcutaneous-adipose tissues were collected from 8 patients. Cells were isolated and cultured. Colony formation was assayed using Colonyze software based on ASTM standards. Cell concentration ([Cell]), CTP concentration ([CTP]) and CTP prevalence (PCTP) were determined. Attributes of culture-expanded cells were compared based on (i) effective proliferation rate and (ii) expression of surface-markers CD73, CD90, CD105, SSEA-4, SSEA-3, SSEA-1/CD15, Cripto-1, E-Cadherin/CD324, Ep-CAM/CD326, CD146, hyaluronan and transcription factors Oct3/4, Sox-2 and Nanog using flow cytometry. RESULTS: Mean [Cell], [CTP] and PCTP were significantly different between MS and TS samples (P = 0.03, P = 0.008 and P= 0.0003), respectively. AT-derived cells generated the highest mean total cell yield at day 6 of culture-4-fold greater than TS and more than 40-fold greater than MS per million cells plated. TS colonies grew with higher mean density than MS colonies (290 ± 11 versus 150 ± 11 cell per mm2; P = 0.0002). Expression of classical-mesenchymal stromal cell (MSC) markers was consistently recorded (>95%) from all tissue sources, whereas all the other markers were highly variable. CONCLUSIONS: The prevalence and biological potential of CTPs are different between patients and tissue sources and lack variation in classical MSC markers. Other markers are more likely to discriminate differences between cell populations in biological performance. Understanding the underlying reasons for variation in the concentration, prevalence, marker expression and biological potential of CTPs between patients and source tissues and determining the means of managing this variation will contribute to the rational development of cell-based clinical diagnostics and targeted cell-based therapies.

Blocking fibrotic signaling in fibroblasts from patients with carpal tunnel syndrome.

Fibrosis of the subsynovial connective tissue (SSCT) in carpal tunnel syndrome (CTS) patients is increasingly recognized as an important aspect of CTS pathophysiology. In this study, we evaluated the effect of blocking profibrotic pathways in fibroblasts from the SSCT in CTS patients. Fibroblasts were stimulated with transforming growth factor ß1 (TGF-ß1), and then treated either with a specific fibrosis pathway inhibitor targeting TGF-ß receptor type 1 (TßRI), platelet-derived growth factor receptor (PDGFR), epidermal growth factor receptor (EGFR), or vascular endothelial growth factor receptor (VEGFR). Fibrosis array and quantitative real-time polymerase chain reaction of fibrotic genes were evaluated. Array gene expression analysis revealed significant down-regulation of multiple fibrotic genes after treatment with TßRI, PDGFR, and VEGFR inhibitors. No array fibrotic genes were significantly down-regulated with EGFR inhibition. Further gene expression analysis of known CTS fibrosis markers collagen type I A2 (Col1), collagen type III A1 (Col3), connective tissue growth factor (CTGF), and SERPINE1 showed significantly down-regulation after TßRI inhibition. In contrast, VEGFR inhibition significantly down-regulated CTGF and SERPINE1, whereas, PDGFR and EGFR inhibition significantly down-regulated Col3. Taken together the inhibition of TßRI appears to be the primary mediator of fibrotic gene expression in fibroblasts from CTS patients. TGF-ß/Smad activity was further evaluated, and as expected inhibition of Smad activity was significantly down-regulated after inhibition of TßRI, but not with PDGFR, VEGFR, or EGFR inhibition. These results indicate that local therapies specifically targeting TGF-ß signaling alone or in combination offer the potential of a novel local antifibrosis therapy for patients with CTS.

ECSIT links TLR and BMP signaling in FOP connective tissue progenitor cells.

Clinical and laboratory observations strongly suggest that the innate immune system induces flare-ups in the setting of dysregulated bone morphogenetic protein (BMP) signaling in fibrodysplasia ossificans progressiva (FOP). In order to investigate the signaling substrates of this hypothesis, we examined toll-like receptor (TLR) activation and bone morphogenetic protein (BMP) signaling in connective tissue progenitor cells (CTPCs) from FOP patients and unaffected individuals. We found that inflammatory stimuli broadly activate TLR expression in FOP CTPCs and that TLR3/TLR4 signaling amplifies BMP pathway signaling through both ligand dependent and independent mechanisms. Importantly, Evolutionarily Conserved Signaling Intermediate in the Toll Pathway (ECSIT) integrates TLR injury signaling with dysregulated BMP pathway signaling in FOP CTPCs. These findings provide novel insight into the cell autonomous integration of injury signals from the innate immune system with dysregulated response signals from the BMP signaling pathway and provide new exploratory targets for therapeutic approaches to blocking the induction and amplification of FOP lesions.

Interacciones epitelio-mesenquimáticas en el desarrollo testicular / Epithelial-mesenchymal transitions in the development of testis

RESUMEN: La espermatogénesis es un proceso continuo que se inicia durante el desarrollo embriofetal. Las relaciones auto, para y yuxtacrinas indican la interdependencia de las células intersticiales (de Leydig) con las células peritubulares (lamina propia) y células sustentaculares (de Sertoli). Ciertos morfógenos son fundamentales en este proceso. Las células sustentaculares son capaces de regular la diferenciación y función de las células peritubulares e intersticiales a través de la producción de IGF1, TGFA, TGFB y DHH. Las células peritubulares son capaces de producir P-Mod-S, regulando la diferenciación de las células sustentaculares, y a través de FGF2 y FGF9 modulan las transiciones epitelio-mesenquimática entre células sustentaculares y mesonefros. También remodelan la membrana basal del condón testicular y regulan la diferenciación y función de las células intersticiales por medio de IGF1, TGFA y TGFB. Las células intersticiales son las reponsables de la producción de testosterona e INSL3, influyendo en la diferenciación sexual masculina. Se plantea que provienen de células mesenquimales del epitelio celómico y mesonefros. Sin embargo, otros autores proponen su origen a partir de células de la cresta neural. Estas influyen a través de mecanismos paracrinos en la proliferación de las células sutentaculares por medio de activina A, teniendo como resultado la expansión del cordón testicular. Las interacciones entre las distintas poblaciones celulares a través de morfógenos inducen una transición epitelio-mesénquima fundamental en la formación y diferenciación de la gónada masculina.

SUMMARY: Spermatogenesis is a continuous process which starts during the embryo-fetal development. Auto, para and juxtacrine relations indicate the interdependence of the interstitial cells (Leydig) with the peritubular cells (lamina propria) and sustentacular cells (Sertoli). Certain morphogens are fundamental in this process. Sustentacular cells are able to regulate differentiation and function and peritubular interstitial cells through production of IGF1, TGFA, TGFB and DHH. Peritubular cells are able to produce P-Mod-S regulating differentiation sustentacular cells and through FGF2 and FGF9 modulate epithelial-mesenchymal transitions between sustentacular cells and mesonephros. They also remodel the basal membrane of the testicular condom and regulate the differentiation and function of the interstitial cells by means of IGF1, TGFA and TGFB. Interstitial cells are responsible for the production of testosterone and INSL3, influencing male sexual differentiation. It is suggested that they come from mesenchymal cells of the coelomic epithelium and mesonephros. However, other authors propose their origin from cells of the neural crest. These influence through paracrine mechanisms proliferation sutentaculares cells by activin A, resulting in the expansion of cord testicular. The interactions between the different cell populations through morphogens induce a fundamental epithelial-mesenchymal transition in the formation and differentiation of the male gonad.

Programmed biomolecule delivery to enable and direct cell migration for connective tissue repair.

Dense connective tissue injuries have limited repair, due to the paucity of cells at the wound site. We hypothesize that decreasing the density of the local extracellular matrix (ECM) in conjunction with releasing chemoattractive signals increases cellularity and tissue formation after injury. Using the knee meniscus as a model system, we query interstitial cell migration in the context of migratory barriers using a novel tissue Boyden chamber and show that a gradient of platelet-derived growth factor-AB (PDGF-AB) expedites migration through native tissue. To implement these signals in situ, we develop nanofibrous scaffolds with distinct fiber fractions that sequentially release active collagenase (to increase ECM porosity) and PDGF-AB (to attract endogenous cells) in a localized and coordinated manner. We show that, when placed into a meniscal defect, the controlled release of collagenase and PDGF-AB increases cellularity at the interface and within the scaffold, as well as integration with the surrounding tissue.