Kisspeptin (Kp-10) is a neuropeptide that binds to GPR54 receptors, exerting several functions mainly in the nervous and reproductive systems of the body. However, its effects and mechanisms of action on the skeletal sys...Kisspeptin (Kp-10) is a neuropeptide that binds to GPR54 receptors, exerting several functions mainly in the nervous and reproductive systems of the body. However, its effects and mechanisms of action on the skeletal system remain poorly understood. This study evaluated the effects of different concentrations of Kp-10 on in vitro osteogenic differentiation of multipotent mesenchymal stromal cells (MSCs) extracted from the bone marrow (BM) of adult Wistar rats. Two-month-old female rats were euthanized to extract BM from long bones to obtain MSCs. Four experimental groups were established in vitro: a control and Kp-10 at concentrations of 0.01, 0.05 and, 0.1 µg/mL. After induction of osteogenic differentiation, cell viability was evaluated using the 3-(4,5-dimethylthiazol-2-yl)- 2,5-diphenyl tetrazolium bromide (MTT) assay, alkaline phosphatase activity, collagen synthesis, percentage of area covered by MSCs/field and mineralized nodules/field, and immunocytochemistry of the GPR54 receptor tests. Furthermore, evaluation of gene transcripts for type I collagen, Runx-2, Bmp-2, bone sialoprotein, osteocalcin and osteopontin was performed using real-time RT-qPCR. It was observed that MSCs expressed GPR54 receptor to which Kp-10 binds during osteogenic differentiation, promoting a negative effect on osteogenic differentiation. This effect was observed at all the Kp-10 concentrations in a concentration-dependent manner, characterized by a decrease in the activity of alkaline phosphatase, collagen synthesis, mineralized nodules, and decreased expression of gene transcripts for type I collagen, osteocalcin, osteopontin, and Runx-2. Thus, Kp-10 inhibits in vitro osteogenic differentiation of MSCs extracted from the BM of adult Wistar rats.
PURPOSE: Lizard regeneration derives from the re-activation of a number of developmental genes after tail amputation. Among genes with the highest expression, as indicated from the transcriptome, is lix1 which functional...PURPOSE: Lizard regeneration derives from the re-activation of a number of developmental genes after tail amputation. Among genes with the highest expression, as indicated from the transcriptome, is lix1 which functional role is not known. METHOD: An antibody that cross-reacts with the lizard Podarcis muralis lix1 has been utilized to detect by immunofluorescence the sites of localization of the protein in the regenerating tail. RESULTS: Lix1-protein is almost exclusively localized in the regenerating spinal cord (ependyma) and nerves growing into the blastema, in sparse blastema cells but is undetectable in other tissues. CONCLUSIONS: Since the spinal cord is essential to stimulate tail regeneration it is hypothesized that the lix1 protein is part of the signaling or growing factors produced from the regenerating spinal cord that are needed for tail regeneration of the lizard tail.
UV-induced corneal damage is a common ocular surface injury that usually leads to corneal lesions causing persistent inflammation. High mobility group box 1 (HMGB1) is identified as an inflammatory alarm in various tissu...UV-induced corneal damage is a common ocular surface injury that usually leads to corneal lesions causing persistent inflammation. High mobility group box 1 (HMGB1) is identified as an inflammatory alarm in various tissue injuries. Here, this study first evaluates the repair effect of the HMGB1-selective inhibitor GLY in UV-induced corneal damage; Secondly, the inhibitory effect of GLY on UV-induced corneal damage induced inflammation and the potential therapeutic mechanism of GLY were studied. GLY effectively attenuates the expression of UV-induced inflammatory factors and HMGB1, TLR/MyD88, NF-κB signaling pathway genes at the mRNA and protein levels. In addition, RT-PCR and Western Blot experiments after knocking down HMGB1 and TLR2/9 genes showed that GLY alleviated corneal inflammation by inhibiting the HMGB1-TLR/MyD88 signaling pathway. The results of this study show that targeting HMGB1-NF-κB by GLY can alleviate the inflammatory response induced by UV induction.
Pterygium is a common eye surface disease with high recurrence and unclear pathogenesis. In current study, RNA sequencing was conducted in 6 pairs of human pterygium and conjunctival tissues, and Matr3 as a novel candida...Pterygium is a common eye surface disease with high recurrence and unclear pathogenesis. In current study, RNA sequencing was conducted in 6 pairs of human pterygium and conjunctival tissues, and Matr3 as a novel candidate gene was significantly reduced in pterygium compared to control tissues. Moreover, immunoprecipitation was performed to pull down MATR3, and WTAP specially interacting with MATR3 in control but not pterygium was identified by mass spectrum. Immunoprecipitation was performed to validate the interaction between MATR3 and WTAP/METTL3/METTL14 complex. (Methylated) RNA immunoprecipitation was performed to further reveal that the binding affinity of WTAP and MATR3 was lost at 3' UTR of RNA molecules of down-regulated genes in pterygium. Overall, we figured out the loss of intercrossing between MATR3 and N6-methyladenosine methyltransferase complex, as well as indicated the potential impact on transcription of target genes in pterygium.
OBJECTIVE: Idiopathic pulmonary fibrosis (IPF) is considered as a chronic interstitial lung disease with underlying mechanism of IPF remaining unclear, while there are no definitive treatment options. In recent years, sc...OBJECTIVE: Idiopathic pulmonary fibrosis (IPF) is considered as a chronic interstitial lung disease with underlying mechanism of IPF remaining unclear, while there are no definitive treatment options. In recent years, scientists have gradually paid attention to the influence of angiogenesis on IPF. Because IPF is a progressive with microvascular remodeling disorder, scientists have postulated that angiogenesis may also be one of the initiating and contributing factors of the disease. Bupleurum is a common natural Chinese herbal medicine with antibacterial, anti-inflammatory, anti-tumor and other pharmacological effects. As the most important active monomer of Bupleurum, Saikosaponin-d (SSd) is a new discovery with anti-pulmonary fibrosis (PF) activity. This study attempts to investigate the role of SSd in the interference of PF through regulation of angiogenesis in IPF through Angiopoietin (Angpt) /Tie receptor 2 (Tie2) pathway. METHODS: Randomly, we allocated C57BL/6 mice into four groups (n = 20 in each group). Afterwards, establishment of IPF model was accomplished via intratracheal administration of bleomycin (BLM, 5 mg/kg), while corresponding drug intervention was given accordingly. On 3rd, 7th, 14th and 28th days after modeling, we performed histopathological examination through staining. Meanwhile, immunohistochemistry (IHC) of PF and the expression of related factors were observed, while Ang/Tie2 pathway was assessed by ELISA with the effect of SSd on angiogenesis related proteins in IPF being explored with IHC and Western Blot technique. RESULTS: Our results showed that SSd could reduce inflammation and PF levels in lung tissue of experimental mice, while levels of angiogenesis-related factors, namely Tie-2, Ang-1 and ANGPT2 (Ang-2), fibrosis- associated factors like Alpha-smooth muscle actin (α-SMA), collagen-I and hydroxyproline in SSd and dexamethasone (DXM) mice were significantly reduced at each time point compared to BLM (p < 0.01). Additionally, we discovered substantial decreased expressions of Ang-1, Ang-2, Tie-2, α-SMA and collagen-I at protein level in SSd and DXM mice at each time point compared to BLM (p < 0.05). Besides, insignificant differences were observed between SSd and DXM groups (p > 0.05). CONCLUSION: This study has demonstrated that SSd could down-regulate the expression of ANG-1, Ang-2 and Tie2 in the Ang/Tie2 pathway, and may reduce lung inflammation and PF in BLM-induced mice via inhibition of angiogenesis.
BACKGROUND: Adipose mesenchymal stem cell (AMSC)-derived exosomes are promising novel factors for wound repair and regeneration. This study aimed to explore the potential roles and underlying mechanisms of specific miRNA...BACKGROUND: Adipose mesenchymal stem cell (AMSC)-derived exosomes are promising novel factors for wound repair and regeneration. This study aimed to explore the potential roles and underlying mechanisms of specific miRNA in wound healing using AMSC-derived exosomes as carriers. METHODS: The expression profiles of GSE197840 were downloaded to screen for differentially expressed miRNAs (DEmiRNAs), and the corresponding genes of the identified miRNAs were predicted. Next, miRNA-mRNA co-expression networks were constructed and the genes in these networks were subjected to functional analysis. miR-223-3p overexpressed AMSCs were then established to isolate exosomes, and the effects of AMSC-derived exosomes carrying miR-223-3p on wound healing and the related potential mechanisms were further investigated in vivo. RESULTS: 35 DEmiRNAs were identified and a co-expression network containing 22 miRNAs and 91 target genes was constructed. Based on the network, miR-223-3p was the hub node and the genes were significantly enriched in 15 GO terms of biological processes and 14 KEGG pathways, including cAMP, PI3K-Akt, cGMP-PKG, neurotrophin signaling pathway, and dopaminergic synapse. Then, miR-223-3p overexpressed AMSCs-derived exosomes were successfully extracted, and miR-223-3p was found to directly bind with MAPK10. In vivo experiments validated that AMSCs-derived exosomal miR-223-3p could promote wound healing, and up-regulated α-SMA, CD31, COL1A1, COL2A1, COL3A1, and down-regulated MAPK10, TNF-α, IL-β, and IL-6. CONCLUSIONS: AMSC-derived exosomal miR-223-3p may accelerate wound healing by targeting MAPK10.
The ability to differentiate into cells of different lineages, such as bone cells, is the principal value of adult mesenchymal stem cells (MSCs), which can be used with the final aim of regenerating damaged tissue. Due t...The ability to differentiate into cells of different lineages, such as bone cells, is the principal value of adult mesenchymal stem cells (MSCs), which can be used with the final aim of regenerating damaged tissue. Due to its potential use and importance in regenerative medicine and tissue engineering, several questions have been raised regarding the molecular mechanisms of MSC differentiation. As one of the crucial mediators in organism development, the transforming growth factor-beta (TGF-β) superfamily directs MSCs' commitment to selecting differentiation pathways. This review aims to give an overview of the current knowledge on the mechanisms of the TGF-β superfamily in MSCs bone differentiation, with additional insight into the mutual regulation of microRNAs and TGF-β in osteogenesis.
PURPOSE: Telocytes (TCs), a novel type of stromal cells found in tissues, induce macrophage differentiation into classically activated macrophages (M1) types and enhance their phagocytic function. The purpose of this stu...PURPOSE: Telocytes (TCs), a novel type of stromal cells found in tissues, induce macrophage differentiation into classically activated macrophages (M1) types and enhance their phagocytic function. The purpose of this study was to investigate the inhibitory effects of TC-induced M1 macrophages on endometriosis (EMs). METHODS: mouse uterine primary TCs and endometrial stromal cells (ESCs) were isolated and identified using double immunofluorescence staining. For the in vitro study, ESCs were treated with TC-induced M1 macrophages, and the vascular endothelial growth factor (VEGF), matrix metalloproteinase 9 (MMP9), and nuclear factor kappa B (NF-κb) genes were identified by quantitative real-time PCR (qRT-PCR) or western blotting (WB). For the in vivo study, an EMs mouse model received TC-conditioned medium (TCM) via abdominal administration, and characterized the inhibitory effects on growth (lesion weight, volume, and pathology), tissue-resident macrophages differentiation by immunostaining, angiogenic capacity (CD31 and VEGF), invasive capacity (MMP9), and NF-κb expression within EMs lesions. RESULTS: immunofluorescent staining showed that uterine TCs expressed CD34+ and vimentin+, whereas ESCs expressed vimentin+ and cytokeratin-. At the cellular level, TC-induced M1 macrophages can significantly inhibit the expression of VEGF and MMP9 in ESCs through WB or qRT-PCR, possibly by suppressing the NF-κb pathway. The in vivo study showed that macrophages switch from the alternatively activated macrophages (M2) in untreated EMs lesions to the M1 subtype after TCM exposure. Thereby, TC-induced M1 macrophages contributed to the inhibition of EMs lesions. More importantly, this effect may be achieved by suppressing the expression of NF-κb to inhibit angiogenesis (CD31 and VEGF) and invasion (MMP9) in the tissue. CONCLUSION: TC-induced M1 macrophages play a prevailing role in suppressing EMs by inhibiting angiogenic and invasive capacity through the NF-κb pathway, which provides a promising therapeutic approach for EMs.
OBJECTIVES: Cholestatic liver diseases are characterized by hepatocellular damage, cholangiocyte proliferation, and progressive fibrosis. Bile duct ligation (BDL) is widely used to resemble liver injuries induced by chol...OBJECTIVES: Cholestatic liver diseases are characterized by hepatocellular damage, cholangiocyte proliferation, and progressive fibrosis. Bile duct ligation (BDL) is widely used to resemble liver injuries induced by cholestasis. Peroxisome proliferator-activated receptor-gamma coactivator 1 alpha (PGC1α) was reported to play a critical role in multiple biological responses. Nevertheless, whether PGC1α is involved in bile acid metabolism and biliary disorders remains unclear. This study aimed to investigate the effect of PGC1α on hepatic responses after cholestatic injury. MATERIALS AND METHODS: Wild-type mice were subjected to BDL or sham surgery for 14 days and human liver specimens from patients with primary biliary cholangitis (PBC) were collected to detect the expression of PGC1α. Hepatic-specific PGC1α knockout mice (HKO) were constructed and subjected to BDL, in which the effects of PGC1α on cholestatic liver injury were demonstrated by biochemical and histopathological assessments, immunoblotting, and metabolomics. RESULTS: The expression of PGC1α was upregulated in the liver of PBC patients and murine models. Both in vivo and in vitro experiments supported the protective effects of PGC1α on cholestasis-induced hepatocyte injury. Infiltrated inflammatory cells after BDL were decreased in HKO mice. Inhibited Wnt/β-Catenin pathway and enhanced Notch signaling promoted transdifferentiation of hepatic progenitor cells (HPC)/ hepatocytes into cholangiocytes, leading to the greater ductular reaction observed in the HKO mice. But bile acids metabolism and mitochondrial function were not affected due to hepatic PGC1α deficiency in cholestasis. CONCLUSIONS: Hepatic-specific deletion of PGC1α regulated liver regeneration by promoting ductular reactions, thereby exerting protective effects against BDL-induced liver injury, which could be a new potential therapeutic target.
Situated in the oral cavity, the rabbit palatine tonsils are part of the mucosal immune system and help to defend the body against foreign pathogens. Expressed as two oval protrusions in the wall of the oropharynx, the r...Situated in the oral cavity, the rabbit palatine tonsils are part of the mucosal immune system and help to defend the body against foreign pathogens. Expressed as two oval protrusions in the wall of the oropharynx, the rabbit palatine tonsils are characterized by excretory ducts and trabeculae. We here compare paraffin embedded and cryosections of the healthy rabbit tonsils. This analysis centers on evaluating the differential outcomes resulting from the application of these fixation methodologies in conjunction with immunohistochemical assays targeting collagen I, collagen III, fibronectin, α-smooth muscle actin (α-SMA), and ki67. Subsequent recommendations are provided based on our findings. Furthermore, we demonstrate the advantage of an antigen retrieval step in immunohistochemical labeling of paraffin sections. Basic classical histological stainings as HE, GT and elastin were also performed. Comparison of different stainings and labelings was furthermore performed in serial sections, showing that adjacent to the excretory ducts, the tonsillar tissue was particularly composed of collagen I and fibronectin, while the vessel walls were predominantly α-SMA positive. Moreover, PAR-2 immunohistochemical staining was performed, where a small fraction of the cells found in the tonsillar connective tissue were PAR-2 positive (probably a subpopulation of mast cells), as well as the lumen of some excretory ducts and trabeculae. Collagen III on the other hand was only weakly expressed in the tonsils. Proliferating ki67 positive cells were rare. This endeavor serves to furnish the scientific community with reference imagery pertinent to researchers opting for the rabbit palatine tonsil model. The diversity of staining techniques employed herein establishes a foundational repository of images, primed for comparative analysis against pathological conditions. Furthermore, these images hold the potential to illustrate inter-species variations. For instance, they can be juxtaposed against murine or rodent tonsils, or even offer insights into the human context.
Topiramate [2,3:4,5-bis-o-(1-methylethylidene) β-D-fructo-pyranose sulfamate; TPM] is one of the most used new-generation antiepileptic drugs. It has been reported to regulate the differentiation of human bone cells. How...Topiramate [2,3:4,5-bis-o-(1-methylethylidene) β-D-fructo-pyranose sulfamate; TPM] is one of the most used new-generation antiepileptic drugs. It has been reported to regulate the differentiation of human bone cells. However, the molecular mechanism of TPM in osteoblast differentiation is not fully elucidated. In the present study, we examined the effect of TPM on osteogenic differentiation of C3H10T1/2, MC3T3-E1, primary mouse calvarial cells, and primary bone marrow stem cells (BMSCs). Primary cells were isolated from mice calvaria and bone marrow respectively. Expression of the osteogenic gene was determined by RT-PCR. The osteogenic protein levels were measured by Western blot analysis. Alkaline phosphatase (ALP) staining experiment was performed to evaluate ALP activity. Alizarin red s (ARS) staining was performed to measure zebrafish caudal fin regeneration. Treatment of TPM up-regulated the osteogenic genes including distal-less homeobox 5 (Dlx5) and runt-related transcription factor 2 (Runx2). In addition, TPM also increased the Dlx5 and Runx2 protein levels, Smad1/5/9 phosphorylation, and alkaline phosphatase (ALP) activity. Furthermore, TPM activated AMPK, and inhibition of AMPK decreased TPM-induced osteogenic differentiation. In the zebrafish model, osteogenic effect of TPM was identified. TPM was increased amputated caudal fin rays of zebrafish. These results demonstrate that TPM enhances osteogenic differentiation via AMPK-mediated Smad1/5/9 phosphorylation.
Since their relatively recent discovery, telocytes (TCs) have been described as peculiar cells strategically positioned in the stromal tissue component of multiple organ systems of the mammalian body including female rep...Since their relatively recent discovery, telocytes (TCs) have been described as peculiar cells strategically positioned in the stromal tissue component of multiple organ systems of the mammalian body including female reproductive organs (i.e., ovary, uterine tube, and uterus). Nevertheless, current knowledge of TCs in the vagina is very limited. The present study was therefore undertaken to investigate the existence and characteristics of TCs in the stromal tissue of human vaginal mucosa by means of immunohistochemistry, immunofluorescence confocal microscopy, and transmission electron microscopy. In the vaginal lamina propria, TCs were first identified by CD34 immunohistochemistry that revealed the presence of CD34 stromal cells arranged in networks, especially around blood vessels. Double immunofluorescence confocal microscopy allowed to precisely distinguish the perivascular networks of CD34 stromal cells lacking CD31 immunoreactivity from adjacent CD31 microvessels. All the perivascular networks of TCs/CD34 stromal cells situated in the vaginal lamina propria coexpressed platelet-derived growth factor receptor α, which strengthened their identification as TCs. Instead, vaginal mucosal TCs were immunophenotypically negative for c-kit/CD117. The ultrastructural examination confirmed the presence of TCs, namely stromal cells with characteristic cytoplasmic processes (i.e., telopodes) forming labyrinthine networks around blood vessels and releasing extracellular vesicles. Together, our morphological findings provide the first comprehensive demonstration that TCs reside in the human vaginal lamina propria, thus paving the way for further investigation of their putative functions in vaginal mucosal homeostasis and pathophysiology.
BACKGROUND: Each eccrine sweat gland (ESG) is a single-tubular structure with a central lumen, and the formation of hollow lumen in the initial solid cell mass is a key developmental process. To date, there are no report...BACKGROUND: Each eccrine sweat gland (ESG) is a single-tubular structure with a central lumen, and the formation of hollow lumen in the initial solid cell mass is a key developmental process. To date, there are no reports on the mechanism of native ESG lumen formation. METHODS: To investigate the lumen morphogenesis and the lumen formation mechanisms of Sprague-Dawley (SD) rat ESGs, SD rat hind-footpads at E20.5, P1-P5, P7, P9, P12, P21, P28 and P56 were obtained. The lumen morphogenesis of ESGs was examined by HE staining and immunofluorescence staining for polarity markers. The possible mechanisms of lumen formation were detected by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) apoptosis assay and autophagy marker LC3B immunofluorescence staining, and further explored by ouabain intervention experiment. RESULTS: In SD rat ESGs, the microlumen was formed at P1, and the small intact lumen with apical-basal polarity appeared at P3. The expression of apical marker F-actin, basal marker Laminin, basolateral marker E-cadherin was consistent with the timing of lumen formation of SD rat ESGs. During rat ESG development, apoptosis and autophagy were not detected. However, inhibition of Na-K-ATPase (NKA) with ouabain resulted in decreased lumen size, although neither the timing of lumen formation nor the expression of polarity proteins was altered. CONCLUSIONS: Epithelial polarity-driven membrane separation but not cavitation regulates lumen formation of SD rat ESGs. NKA-regulated fluid accumulation drives lumen expansion.
In maize, immunoprecipitation assays have shown that CycD2;2 interacts with KRPs. However, evidence on CycD2;2 or KRPs localization and their possible interaction in specific tissues is lacking and its physiological cons...In maize, immunoprecipitation assays have shown that CycD2;2 interacts with KRPs. However, evidence on CycD2;2 or KRPs localization and their possible interaction in specific tissues is lacking and its physiological consequence is still unknown. This work explores the spatiotemporal presence of CyclinD2s and KRPs, cell cycle regulators, during maize seed germination (18 and 36 h) after soaking on glucose or sucrose (120 mM). CyclinD2s are positive actors driving proliferation; KRPs are inhibitors of the main kinase controlling proliferation (a negative signal that slows down the cell cycle). Cell cycle proteins were analyzed by immunolocalization on longitudinal sections of maize embryo axis in seven different tissues or zones (with different proliferation or differentiation potential) and in the nucleus of their cells. Results showed a prevalence of these cell cycle proteins on embryo axes from dry seeds, particularly, their accumulation in nuclei of radicle cells. The absence of sugar caused the accumulation of these regulators in different proliferating zones. CyclinD2 abundance was reduced during germination in the presence of sucrose along the embryo axis, while there was an increase at 36 h on glucose. KRP proteins showed a slight increase at 18 h and a decrease at 36 h on both sugars. There was no correlation between cell cycle regulators/DNA co-localization on both sugars. Results suggest glucose induced a specific accumulation of each cell cycle regulator depending on the proliferation zone as well as nuclear localization which may reflect the differential morphogenetic program regarding the proliferation potential in each zone, while sucrose has a mild influence on both cell cycle proteins accumulation during germination. Whenever CycD2s were present in the nucleus, KRPs were absent after treatment with either sugar and at the two imbibition times analyzed, along the different embryo axe zones.
Central giant cell granuloma (CGCG) is a benign jaw lesion with variable clinical behavior. Cell cannibalism is a cellular process associated with aggressiveness and invasion in malignant neoplasms. Here, we morphologica...Central giant cell granuloma (CGCG) is a benign jaw lesion with variable clinical behavior. Cell cannibalism is a cellular process associated with aggressiveness and invasion in malignant neoplasms. Here, we morphologically investigated cell cannibalism as an auxiliary method to predict CGCG clinical behavior. Cell cannibalism was quantitatively evaluated in 19 cases of peripheral giant cell granuloma (PGCG), 38 cases of CGCG (non-aggressive and aggressive), and 19 cases of giant cell tumor of bone (GCT) stained with hematoxylin and eosin. T-test was performed to assess the differences between the variables analyzed (p ≤ 0.05). Cell cannibalism was identified in 21% of non-aggressive CGCGs and 68.4% of aggressive CGCGs. A significantly higher amount of cannibal multinucleated giant cells (CMGC) was observed in aggressive CGCG compared to PGCG and non-aggressive CGCG (p = 0.042; p = 0.044, respectively). There were no significant differences in the CMGC index between non-aggressive CGCG and PGCG (p = 0.858) and between aggressive CGCG and GCT (p = 0.069). CGGC cases that exhibited rapid growth and tooth displacement and/or root resorption had a higher amount of CMGC (p = 0.035; p = 0.041, respectively). Cell cannibalism can be identified in CGCG through routine anatomopathological examination. The quantification of CMGC can help to predict the clinical behavior of central giant cell granuloma.
BACKGROUND: Fibroblasts (FBs) have been widely used as a typical in vitro cell model for investigating the biological processes and cell pathophysiological mechanisms. However, FBs are prone to senescence in cell culture...BACKGROUND: Fibroblasts (FBs) have been widely used as a typical in vitro cell model for investigating the biological processes and cell pathophysiological mechanisms. However, FBs are prone to senescence in cell culture process after several passages. Thus, a new approach to cell culture is quite required to enhance the viability of cells. OBJECTIVE: To explore a novel method of cell culture based on skin FBs. METHODS: Dermal tissue blocks were obtained from BALB/c neonatal mice and randomly divided into experimental group and control group. The experimental group received the newly improved culture method, namely, continuous adherence subculture of tissue block (CASTB) method; while the traditional subculture method was applied in the control group. Cells at 1st, 5th and 10th passages were collected and identified by using histological/immunohistochemical and western blot analysis. Cellular viability, proliferation, senescence and apoptosis were analyzed through application of cell growth curve, CCK-8 assay, Ki67 assay, PCNA protein analysis, β-galactosidase staining, flow cytometry and western blot analysis. RESULTS: Cells under two culture patterns exhibited spindle/irregular shape and vimentin positive expression. With the increase of passage times, the cellular growth rate in the control group gradually decreased, but no alterations emerged from the experimental group. CASTB method remarkably promoted cell growth and proliferation. Moreover, a greatly lower apoptosis and senescence tendency appeared in the experimental group than the control group with passages increasing. CONCLUSION: The method of CASTB is superior to traditional subculture, offering a large number of primary FBs with higher efficiency and success rate and being worth of further popularization and application.
The vomeronasal organ is an olfactory organ found in amphibians and higher vertebrates. Type 1 vomeronasal receptors, one of the major olfactory receptors in vertebrates, are expressed in the vomeronasal organ in mammals...The vomeronasal organ is an olfactory organ found in amphibians and higher vertebrates. Type 1 vomeronasal receptors, one of the major olfactory receptors in vertebrates, are expressed in the vomeronasal organ in mammals. In amphibians and fish, they are expressed in the olfactory epithelium. The lungfish, which is the species of fish most closely related to amphibians, has a primitive vomeronasal organ: the recess epithelium. Expression of type 1 vomeronasal receptors has been reported in both the olfactory epithelium and the recess epithelium in three species of African lungfish and one species of South American lungfish. However, a previous study suggested that in the African lungfish Protopterus dolloi these receptors are expressed only in the olfactory epithelium. In this study, we identified 21 type 1 vomeronasal receptor genes in P. dolloi and examined the expression sites in the olfactory organ. In P. dolloi, most cells expressing the type 1 vomeronasal receptor were distributed in the olfactory epithelium, but a few were also found in the recess epithelium. This implies that the functions of the olfactory epithelium and the primitive vomeronasal organ are incompletely separated, and that all extant African and South American lungfish share this trait.
HSCs (hepatic stellate cells) contribute to the excessive extracellular matrix (ECM) deposition plays a key role in the progression of hepatic fibrosis. The present study focused on the hepatoprotective effect of Ginsenos...HSCs (hepatic stellate cells) contribute to the excessive extracellular matrix (ECM) deposition plays a key role in the progression of hepatic fibrosis. The present study focused on the hepatoprotective effect of Ginsenoside Rc (Rc), one of the protopanaxadiol type ginsenoside, which has contributed to reverse activated HSCs to improve hepatic fibrosis via regulating Nur77-TLR4/MyD88 signaling pathway. We established the hepatic fibrosis model by intraperitoneal injection of carbon tetrachloride (CCl). And HSCs were stimulated with TGF-β, followed by silencing of Nur77, and then incubated in Rc. Rc significantly alleviated histopathological changes, reduced serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels. Rc could upregulate the Nur77 and downregulate fibrosis markers in the liver of mice, including decreasing the expressions of α-SMA, Collagen-I, the ratio of TIMP-1/MMP-13. Rc significantly increased the expression of Nur77 and suppressed the production of ECM in HSCs. Rc inhibited TLR4 signaling pathway, consequently reversing the inflammatory response, including the production of MyD88, IRAK1, IRAK4 and IL-23. When Nur77 was knocked in TGF-β-stimulated HSCs, TLR4 and α-SMA production were increased. Rc suppressed these activatory effects in Nur77 knockdown HSCs. Rc reduced inflammatory reaction by regulating the Nur77-TLR4 signaling pathway while suppressing the fibrogenesis suggesting, underscoring a promising approach of Rc for the treatment in hepatic fibrosis. Targeting Nur77-TLR4 signaling in HSCs would be the potential strategy for Rc against hepatic fibrosis.
Even after more than two years of intensive research, not all of the pathophysiological processes of Coronavirus Disease 2019 (COVID-19), induced by severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2) infec...Even after more than two years of intensive research, not all of the pathophysiological processes of Coronavirus Disease 2019 (COVID-19), induced by severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2) infection, have been fully elucidated. The initial virus-host interaction at the respiratory epithelium plays a crucial role in the course and progression of the infection, and is highly dependent on the glycosylation pattern of the host cell and of the secreted mucins. Glycans are polysaccharides that can be attached to proteins and thereby add to their stability and functionality. Lectins are glycan-binding proteins that recognize specific glycan motifs, and lectin histochemistry is a suitable tool to visualize and examine glycosylation pattern changes in tissues. In this study we used lectins with different glycan-specificities for the visualization of glycosylation pattern changes in the respiratory tract of SARS-CoV-2 infected Golden Syrian hamsters. While some lectins (LEL, STL) enable the visualization of the damage to alveolar type 1 pneumocytes, other lectins, e.g., GSLI, visualized the loss and subsequent hyperplasia of type 2 pneumocytes. UEAI staining was co-localized with KI67, a proliferation marker. Double staining of lectins LEL, STL and WGA with specific immune cell markers (Iba1, CD68) showed co-localization and the dominant infiltration of monocyte-derived macrophages into infected alveolar tissue. The elucidation of the glycosylation pattern of the respiratory tract cells in uninfected and infected Golden Syrian hamsters revealed physiological and pathological aspects of the disease that may open new possibilities for therapeutic development.