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Background context Hypertrophy of the ligamentum flavum (LF) is a major contributor to the development of lumbar spinal canal stenosis (LSS). Although previous studies have identified some factors related to hypertrophy of the LF, the etiology remains unclear. It is well known that myofibroblasts have a key role in the pathology of fibrosis in other tissues, including the skin, liver, kidney, and lung. We hypothesized that myofibroblasts were also important players in the pathology of fibrosis in the LF. Purpose To elucidate the distribution and role of myofibroblasts in the hypertrophic LF. Study design A histological, immunohistochemical, and gene expression analysis of the LF in the human lumbar spine. Patient sample Hypertrophic LF tissue samples were collected from patients with LSS. Outcome Measures Histology, immunohistochemistry, microarray, reverse transcription-quantitative polymerase chain reaction, western blotting and enzyme-linked immunosorbent assay. Methods The degree of fibrosis in the dural and dorsal layers of the LF was evaluated by Masson's trichrome tissue staining. Collagen gene expression was evaluated by quantitative reverse transcription polymerase chain reaction. Immunostaining of αSMA was performed to evaluate localization of myofibroblasts in LF tissue. The association between gene expression of alpha-smooth muscle actin (αSMA) and that of several types of collagen was investigated. The signal activated on the dorsal side of LF was examined by gene set enrichment analysis using microarray data. Expression levels of αSMA and several types of collagen in LF fibroblasts were investigated under hypoxic conditions. Results In the histological study using Masson's trichrome staining, the fibrosis score was significantly higher in the dorsal layer than in the dural layer. Gene expression levels for several types of collagen (COL1A1, COL1A2, COL3A1, COL5A1, COL6A1, and COL11A1) and heat shock protein 47 (a collagen-specific chaperone) were significantly higher in the dorsal layer. Furthermore, immunohistochemistry revealed a significantly greater number of αSMA-stained cells in the dorsal layer. There was a strong correlation of αSMA mRNA expression with COL1A-1 in LF fibroblasts. Gene set enrichment analysis showed that the set of fibrosis-related gene signals, including those for epithelial-mesenchymal transition, hypoxia, and inflammation, were significantly upregulated in the dorsal layer compared with the dural layer. Under hypoxic stimulation, expression of αSMA and several types of collagen was increased in LF fibroblasts. Conclusions This study is the first to reveal that myofibroblast expression levels are higher in the dorsal layer of the LF than in the dural layer. We confirmed that hypertrophy of the LF in LSS is associated with increased expression of myofibroblasts in the dorsal layer. Hypoxia could be a cause of expression of myofibroblasts leading to fibrosis and finally to hypertrophy of the LF. Clinical significance The results of this study partially elucidate the molecular mechanisms of LF hypertrophy and suggest that myofibroblasts may be involved in age-related degeneration of the LF.

Introduction A challenge in phonosurgery is achieving good vocal quality with minimal vocal fold fibrosis. Fibrin glue can be applied to minimize fibrosis; however, its use in the larynx is based primarily on clinical experience, particularly in extensive lesions. Objective The objective of this study was to evaluate the effect of application of fibrin glue on collagen concentration at the late phase of the healing process after detaching a pedicled flap of the vocal fold cover in rabbits. Methods In this prospective animal study, twelve adult male rabbits underwent laryngeal microsurgery, in which an incision was made along the entire length of both vocal folds, followed by extensive mucosal detachment and section of the incision ends. Fibrin glue was applied in the left vocal fold, and the mucosa was repositioned. In the right vocal fold, the mucosa was repositioned without treatment with fibrin glue. After 3-months, the rabbits were euthanized. Histological analyses were performed, and the data collected were subjected to statistical analysis. Results Vocal folds treated with fibrin glue presented higher collagen concentration in Masson trichrome staining and significantly higher (p< 0.05) collagen concentration in picrosirius red staining compared to control vocal folds. Conclusion Treatment with fibrin glue led to greater vocal fold fibrogenesis in the present study. Nonetheless, further studies are necessary to determine the prophylactic effect of sealants in laryngeal surgeries that require extensive detachment of the vocal fold cover. Resumo Introdução Um desafio na fonocirurgia é conseguir uma boa qualidade vocal com o mínimo de fibrose das pregas vocais. A cola de fibrina pode ser aplicada para minimizar a fibrose; porém, seu uso na laringe é baseado principalmente na experiência clínica, particularmente em lesões extensas. Objetivo Avaliar o efeito da aplicação da cola de fibrina na concentração de colágeno na fase tardia do processo de cicatrização, após o descolamento do retalho pediculado da cobertura da prega vocal em coelhos. Método Neste estudo prospectivo com animais, 12 coelhos adultos machos foram submetidos à microcirurgia de laringe, na qual uma incisão foi feita em toda a extensão de ambas as pregas vocais, seguida de extenso descolamento da mucosa e secção das extremidades da incisão. A cola de fibrina foi aplicada na prega vocal esquerda e a mucosa foi reposicionada. Na prega vocal direita, a mucosa foi reposicionada sem tratamento com cola de fibrina. Após três meses, os coelhos foram sacrificados. Análises histológicas foram feitas e os dados coletados foram submetidos à análise estatística. Resultados As pregas vocais tratadas com cola de fibrina apresentaram maior concentração de colágeno na coloração tricrômica de Masson e concentração significativamente maior de colágeno (p < 0,05) na coloração com picrosirius red comparadas às pregas vocais controle. Conclusão O tratamento com cola de fibrina resultou em maior fibrogênese das pregas vocais no presente estudo. Contudo, novos estudos são necessários para determinar o efeito profilático dos selantes em cirurgias de laringe que requerem extenso descolamento da cobertura das pregas vocais.

The liver is the largest gland of the gastrointestinal tract having both exocrine and endocrine functions. Developmentally it arises as a ventral outgrowth from the gut endoderm during 3rd week of intrauterine life. The foetal liver is very important because of its synthetic and hemopoietic potential. The present work aimed to see the detailed histogenesis and development of the foetal liver by cytological, immunohistochemical and ultrastructural study. The liver tissue of nine aborted foetuses of various gestational age were studied. For cytology: special stains like Masson trichrome, periodic acid Schiff and reticulin were used, immunohistochemical staining was performed with triple antibodies (c-myc, Ki-67 and Ber-H2), and for ultrastructure: aluminium mounted specimens were coated with gold and argon gas and observed under scanning electron microscopy (EM). Cytology and immunohistochemistry showed the development of duct patterns and hemopoietic patterns in all stages of fetogenesis. The ductal plate was marked by the layer of dark brown staining cells at the edge of two portal tracts. The haemopoietic cells with sinusoids and aggregation of hepatocytes were observed in the early weeks of gestation. EM showed tree-like branching of a portal canal depicting hepatic segmentation of foetal liver. The organizational changes in lobular pattern, duct pattern, and microstructure of liver during fetogenesis are very crucial to achieve the adult morphology in feature. Histogenesis of the foetal liver follows a multistep process depending upon the gestational age, any deviation from normalcy may lead to structural and functional abnormality later in life.

Background Histologic analyses of deep vein thrombi (DVTs) have used autopsy samples and animal models. To the best of our knowledge, no previous study has reported on thrombus composition after percutaneous mechanical extraction. Because elements of chronicity and organization render thrombus resistant to anticoagulation and thrombolysis, a better understanding of clot evolution could inform therapy. Methods We performed a histologic evaluation of DVTs from consecutive patients who had undergone mechanical thrombectomy for extensive iliofemoral DVTs using the Clottriever/Flowtriever device (Inari Medical, Irvine, Calif). The DVTs were scored using a semiquantitative method according to the degree of fibrosis (collagen deposition on trichrome staining) and organization (endothelial growth with capillaries and fibroblastic penetration). Results Twenty-three specimens were available for analysis, with 20 presenting as acute DVT (≤14 days from symptom onset). Of the 23 patients, 11 (48%) had had >5% fibrosis (ie, collagen deposition) and 14 (61%) had had >5% organization (ie, endothelial growth, capillaries, fibroblasts). Four patients with acute DVT had had ≥25% organized thrombus and two had had ≥25% collagen deposition. Of the 20 patients with acute DVT, 40% had had >5% fibrosis and 55% had had >5% organization. The acuity of DVT did not correlate with the amount of fibrosis or organizing scores. Conclusions A large proportion of patients with acute DVT will have histologic elements of chronicity and fibrosis. A better understanding of the relationship between such elements and the response to anticoagulant agents and fibrinolytic drugs could inform our approach to therapy.

BACKGROUND Pregnant women with a Fontan circulation have a high risk of obstetric complications, such as preterm delivery and small for gestational age (SGA), which may be affected by low blood flow to the placenta and hypoxia. This study investigated placental pathology in a Fontan circulation.Methods and Results:Eighteen pregnancies in 11 women with a Fontan circulation were reviewed. Pregnancy outcomes showed 9 miscarriages and 9 live births, with 4 preterm deliveries. Five neonates were SGA (

Objective To present the pathologic analysis of female urethral strictures obtained during reconstructive urethroplasty. Methods Nine separate female urethral tissue specimens were obtained during dorsal vaginal graft urethroplasty by a single surgeon (S.P.P.). Samples were serially sectioned and fixed in 10% formalin 6 to 12 hours before routine processing in paraffin blocks. Serial 5-µm sections were subjected to H&E, Masson trichrome, and elastin staining. End point analysis included evaluation for epithelial hyperplasia and cell type, mucosal edema, degree of fibroblast/inflammatory cell infiltrate, and elastin fiber density and distribution. Results Nine specimens were examined. Six specimens had epithelial linings of stratified squamous epithelium overlying fibrosis (67%), 1 had mixed squamous and urothelial epithelium, and 2 had only urothelial epithelium. Two specimens (29%) showed acute injury with prominent squamous papillary hyperplasia, focal erosion, and patchy mucosal hemorrhage. Areas of urethral stricture were variably thickened, with increased, densely packed collagen fibers and associated mucosal lymphocytic inflammation ranging from mild and patchy to focally dense with lymphoid aggregates. The highest elastin fiber density appeared to be associated with vessels and overlying muscle bundles in the submucosa. Conclusions Further elucidation of histopathologic characteristics may illuminate more appropriate therapeutic pathways for female urethral stricture disease management.

This study estimated the effect of exposure to propiconazole through implementation and residues in finishing pigs. We analyzed the expression of fibrosis-related genes and performed histological analysis of the blood, liver, kidney, muscle, ileum, and fat tissues. The animals were exposed for 28 d to different concentrations of propiconazole (0.09, 0.44, 0.88, 4.41, and 8.82 mg/kg bw/d). Quantitative, gene expression, and histological analyses in tissues were performed using liquid chromatography mass spectrometry, real-time PCR, and Masson’s trichrome staining, respectively. Final body weight did not differ among groups. However, genes involved in fibrosis were significantly differentially regulated in response to propiconazole concentrations. Glucose, alanine aminotransferase, and total bilirubin levels were significantly increased compared with those in the control group, while alkaline phosphatase level was decreased (p

INTRODUCTION Studying the pathological changes of ligaments in patients with haemophilic arthritis (HA) has important significance for guiding the release of ligaments during total knee arthroplasty (TKA) and exploring interventions to prevent ligament lesions. AIM This study was conducted to show the pathological changes and investigate the lysine oxidase (LOX) and matrix metalloproteinase (MMP)-1, -2, and -3 levels in the ligaments of patients with HA compared with those of patients with osteoarthritis (OA). METHODS Ligaments obtained during the TKA were stained with Masson trichrome, Verhoeff-Van Gieson and haematoxylin and eosin to show the basic pathological changes. Collagen I, elastin, LOXs and MMP-1, -2, and -3 expression levels were detected via western blot. LOX and MMP-1, -2, and -3 mRNA expression levels were analysed via quantitative real-time PCR. RESULTS Compared with OA ligaments, HA ligaments were constructed more loosely with wider gaps, more breaks, haemocytodeposition and local hypertrophy among the fibres. LOXs and MMP mRNA expression levels were upregulated in the HA tissues, which was consistent with the western blot results. Collagen I and elastin levels were also higher in patients with HA. CONCLUSIONS The metabolism of the ligaments in patients with HA is more complex than in those with OA, and the ligaments of patients with HA have stronger healing and destruction processes. This pathology is related to iron overload and imbalanced inflammatory factors due to repeated intra-articular bleeding.

Osteomyelitis associated with periprosthetic joint infection (PJI) signals a chronic infection and the need for revision surgery. An osteomyelitic bone exhibits distinct morphological features, including evidence for osteolysis and an accelerated bone remodelling into poorly organised, poor-quality bone. In addition to immune cells, various bone cell-types have been implicated in the pathology. The present study sought to determine the types of bone-cell activities in human PJI bones. Acetabular biopsies from peri-implant bone from patients undergoing revision total hip replacement (THR) for chronic PJI (with several identified pathogens) as well as control bone from the same patients and from patients undergoing primary THR were analysed. Histological analysis confirmed that PJI bone presented increased osteoclastic activity compared to control bone. Analysis of osteocyte parameters showed no differences in osteocyte lacunar area between the acetabular bone taken from PJI patients or primary THR controls. Analysis of bone matrix composition using Masson’s trichrome staining and second-harmonic generation microscopy revealed widespread lack of mature collagen, commonly surrounding osteocytes, in PJI bone. Increased expression of known collagenases, such as matrix metallopeptidase (MMP) 13, MMP1 and cathepsin K (CTSK), was measured in infected bone compared to non-infected bone. Human bone and cultured osteocyte-like cells experimentally exposed to Staphylococcus aureus exhibited strongly upregulated expression of MMP1, MMP3 and MMP13 compared to non-exposed controls. In conclusion, the study identified previously unrecognised bone-matrix changes in PJI caused by multiple organisms deriving from osteocytes. Histological examination of bone collagen composition may provide a useful adjunct diagnostic measure of PJI.

Mesenchymal stem cells (MSCs) have become a promising therapeutic strategy for scleroderma. Exosomes derived from MSCs (MSC-exosomes) possess functional properties similar to those of their source cells. In this study, we aimed to explore the potential role of MSC-exosomes in the treatment of scleroderma. MSC-exosomes were isolated from human umbilical cords through ultracentrifugation and characterized. An experimental fibrosis model was established in BALB/c mice by a subcutaneous injection of bleomycin, followed by treatment with MSC-exosomes or MSC infusions once a week for a total of four doses. Using hematoxylin and eosin and Masson's trichrome staining and immunohistochemistry, hydroxyproline content, and quantitative real-time polymerase chain reaction analyses, we investigated the effects of MSC-exosomes on dermal fibrosis and explored the underlying mechanism. MSC-exosome treatment restored the dermal architecture, reduced dermal thickness, and partially increased subcutaneous adipose tissue thickness. In addition, MSC-exosomes inhibited the expression of collagen (COL)-I, COL-III, and α-smooth muscle actin. The transforming growth factor (TGF)-β/Smad signaling pathway was also suppressed in MSC-exosome-treated mice. Taken together, our results suggest that MSC-exosomes can attenuate myofibroblast activation and collagen deposition in dermal fibrosis by downregulating the TGF-β/Smad signaling pathway. Therefore, the use of MSC-exosomes may be a potential therapeutic approach for the treatment of scleroderma.

Purpose To evaluate the biomechanical and histologic effects on Achilles tendon repair of inhaled combusted tobacco versus nicotine exposure via electronic cigarette versus a control group in a small-animal model (Sprague-Dawley rat). Methods Fifty-four Sprague-Dawley rats were randomized into 3 groups: combusted tobacco, e-cigarettes, or control. Experimental rats were exposed to research cigarettes or e-cigarette vapor in a smoking chamber for 4 weeks. Surgical transection and repair of the Achilles tendon were then completed, followed by 2 additional weeks of exposure. Achilles tendons were harvested, and biomechanical tensile testing was performed. Histologic evaluation was completed, including hematoxylin-eosin staining, trichrome staining, and immunohistochemistry analysis for type I and type III collagen. Results The control group showed the highest mean tensile load to failure, at 41.0 ± 10.4 N (range, 18.3-55.1 N); the cigarette cohort had the second highest mean, at 37.3 ± 11.1 N (range, 14.0-54.7 N); and finally, the vaping group had the lowest mean, at 32.3 ± 8.4 N (range, 17.8-45.1 N). One-way analysis of variance showed a significant difference in load to failure when comparing the control group with the e-cigarette group (P = .026). No statistical difference was detected between the control group and cigarette group (P = .35) or between the e-cigarette group and cigarette group (P = .23). Stiffness and qualitative histologic analysis showed no difference among groups. Conclusions This investigation shows that in a rat model, nicotine exposure via e-cigarette significantly impedes the biomechanical healing properties of Achilles tendon surgical repair. Clinical Relevance The results indicate that although e-cigarettes are often used as a perceived “safer” alternative to smoking, their use may have a detrimental effect on tendon load to failure.

The purpose of this study was to determine the efficacy of highly concentrated tricalcium phosphate (TCP) biocomposite screws on local bone formation in a rabbit model of osteoporosis induced by bilateral ovariohysterectomy (OHE). Fourteen 24-week-old female New Zealand rabbits (weight, 3-3.5 kg) were divided into two groups: 1) OHE and biodegradable poly(lactic-co-glycolic acid) (PLGA) without s-TCP plate or screw insertion (OHE/BSR) group; and 2) OHE and biocomposite PLGA with highly concentrated s-TCP plate and screw insertion (OHE/HCCP). Both groups underwent bilateral OHE and had two different types of screws and plates inserted at the proximal tibia. Bilateral tibiae were extracted at 25 weeks post-OHE. The extracted tibiae were scanned with ex-vivo micro-computed tomography (micro-CT). Parameters including bone mineral density (BMD), trabecular bone volume (BV/TV), trabecular number (Tb.N), trabecular thickness (Tb.Th), and trabecular separation (Tb.Sp) were evaluated after staining the tibial samples with hematoxylin and eosin (H&E) and Masson's trichrome. We then performed pathological assessments. Micro-CT images revealed improved new bone formation near the implant in the OHE/HCCP group with higher values of BMD, BV/TV, and Tb.N but lower values of Tb.Th and Tb.Sp than the OHE/BSR group. Analyses of H&E and Masson's trichrome staining showed better new bone formation around the implant in the OHE/HCCP group than in the OHE/BSR group. The use of highly concentrated TCP biocomposite screw and plate might improve local bone formation and facilitate osteoconductivity in an osteoporotic rabbit model. This article is protected by copyright. All rights reserved.

Objectives The first goal of this study was to synthesize the silver nanoparticles Alcaligenes xylosoxidans exopolysaccharide (Ag-AXEPS). The second objective was to analyse the role of Ag-AXEPS nanoparticles (NPS) in treating bleomycin (BLM)-induced lung fibrosis. Methods Intratracheal bleomycin (2.5 U/kg) was administered to prompt pulmonary fibrosis in rats, and pulmonary fibrosis was treated with Ag-AXEPS nanoparticles (100 ppm/twice a week for four weeks). Key findings Ag-AXEPS nanoparticles significantly decreased the diversity of pulmonary inflammatory agents in rats with BLM-induced fibrosis. Reduced levels of respiratory tumor necrosis factor-alpha, monocyte chemotactic protein-1, matrix metalloproteinases (MMP-2 and MMP-9) were observed on treatment with synthesized Ag-AXEPS. Similarly, the treatment decreased IL-12, mRNA levels of BAX and plasma fibrosis markers like N-terminal procollagen III propeptide and transforming growth factor-β1. On the other hand, the treatment increased mRNA BCL2 and total antioxidant capacity. It also lowered the level of fibrosis, as was shown by a quantified pathologic study of hematoxylin–eosin-stained lung parts. The treatment, however, ensured that lung collagen was restored, as assessed by Masson’s trichrome stain, and that overall survival was increased and enhanced. Conclusions Our work showed that nanoparticles could be obtained at 37°C and may be a possible pulmonary fibrosis therapeutic agent.

Objectives We aimed to investigate the effect of sugammadex on the motor, sensory and deep sensory block in the sciatic nerve created by bupivacaine in rats. Materials and methods 18 Sprague-Dawley adult male rats treated with unilateral sciatic nerve block by bupivacaine (0.2 ml) were randomly divided into three groups. Control group (Group C, n = 6, 1.5 mL saline) perineural sugammadex group (Group PNS, n = 6, 16 mg/kg) and intraperitoneal sugammadex group (Group IPS, n = 6, 16 mg/kg) Motor, sensory, and deep sensory functions were evaluated every 10 minutes by a blind researcher. 6 tissue samples each belonging to the sciatic nerve, 1.5 cm in length and 0.2 cm in diameter, were taken from paraffin blocks. Sections of 3-4 micrometers were stained with Hematoxylin + Eosin, Masson Trichrome dyes and examined under a light microscope. Results There was no statistically significant difference between 3 groups in terms of the time to return to normal motor, sensory and deep sensory function. There was also no significant difference in edema, extracellular matrix, and myelin. Inflammatory cells were seen in all groups, mainly epineurium, epineurium, and perineurium. Conclusion There are findings of no histological effects or effects on local block of sugammadex in rats undergoing sciatic nerve block.

Background. Hypertrophic scar (HS) is a benign fibroproliferative skin disease resulting from an aberrant wound healing process and can cause aesthetic and functional damage to patients. Currently, there is no ideal treatment to treat this disease. Galangin, a natural active bioflavonoid compound, is suggested to inhibit fibrosis and proliferation in certain cells. Methods. In this study, we found Galangin could attenuate abnormal scar formation in an HS rabbit ear model. Additionally, the HE staining shows Galangin reduced scar elevation index (SEI) and Masson’s trichrome staining changed collagen deposition. Results. The expressions of type I collagen, type III collagen, and TGF-β1 were much lower under a proper dose of Galangin treatment, and Smad7 expression was also enhanced, which are examined by real-time PCR, immunohistochemistry, and western blot. Conclusion. Our data indicated that Galangin can alleviate dermal scarring via the TGF-β/Smad signaling pathway probably by upregulating Smad 7 expression and, thus, suppressing the expression of type I and type III collagens and TGF-β1.

Research question Can preoperative or perioperative intervention reduce the risk of recurrence of endometriosis caused by either incomplete excision or spillage and dissemination? Design A mouse model of endometriosis recurrence caused by spillage and dissemination was first established using 24 female Balb/c mice. The spillage and dissemination model was used to test the efficacy of preoperative use of ketorolac, perioperative use of aprepitant and combined use of propranolol and andrographolide in a prospective, randomized mouse experiment involving 75 mice. The efficacy of these preoperative and perioperative interventions in a mouse recurrence model caused by incomplete excision was also tested using 72 mice. In all experiments, the baseline body weight and hotplate latency of all mice were measured and recorded before the induction of endometriosis, before the primary surgery and before sacrifice. In addition, all lesions were excised, weighed and processed for quantification and immunohistochemistry analysis of E-cadherin, α-SMA, VEGF, ADRB2 and putative markers of recurrence PR-B, p-p65, as well as Masson trichrome staining. Results All interventions substantially and significantly suppressed the outgrowth of endometriotic lesions and reduced the risk of recurrence caused by either spillage and dissemination or incomplete excision (P = 0.0007 to 0.042). These interventions also significantly attenuated the generalized hyperalgesia, inhibited the staining of α-SMA, p-p65, VEGF and ADRB2 but increased staining of E-cadherin and PR-B, resulting in reduced fibrosis. Conclusion Given the excellent safety profiles of these drugs, these data strongly suggest that preoperative and perioperative intervention may potentially reduce the risk of endometriosis recurrence effectively.

Utilization of functional-protective coatings for implants based on corundum ceramics seems promising from the point of view of stability, bioinertness, and low cost. In order to study the histo­logical criteria for evaluating the osteoinductive properties of functional protective coatings, 6 types of coatings were studied on an experimental animal model: 90 Wistar rats were implanted with 6 test types of implants with various combinations of surface treatment (sand-blasting, surface treatment with a plasma torch with simultaneous application of aluminum oxide, powdered titanium, etc.) with an exposure of 1, 2 and 4 weeks. After euthanasia, a histological exa­mination of decalcified bone with Masson-Goldner trichrome staining and TRAP-histochemical reaction for osteoclasts was performed. The obtained results de­monstrated significantly higher osteoinductive properties of functional protective coatings with a more pronounced roughness (Ra>10 μm) compared to an untreated titanium surface after 2 (p

Background:. The effectiveness of bronchial thermoplasty (BT) has been reported in patients with severe asthma. This study compared the effects of BT and cryoballoon ablation (CBA) therapy on the airway smooth muscle (ASM). Methods:. Eight healthy male beagle dogs were included in this experiment. In the preliminary experiment, one dog received BT treatment for both lower lobe bronchus, another dog received CBA treatment for 7 s on the upper and lower lobe of right bronchus, and 30 s on the left upper and lower lobe. The treatments were performed twice at an interval of 1 month. In subsequent experiments, the right lower lobe bronchus was treated with BT, and the left lower lobe bronchus was treated with CBA. The effects of treatment were observed after 1 (n = 3) month and 6 months (n = 3). Hematoxylin-eosin staining, Masson trichrome staining, and immunohistochemical staining were used to compare the effects of BT and CBA therapy on the ASM thickness, collagen fibers synthesis, and M3 receptor expression after treatment. One-way analysis of variance with Dunnett post hoc test was used to analyze the differences among groups. Results:. In the preliminary experiment, the ASM ablation effect of 30-s CBA was equivalent to that of 7-s CBA (ASM thickness: 30.52 ± 7.75 μm vs. 17.57 ± 15.20 μm, P = 0.128), but the bronchial mucociliary epithelium did not recover, and large numbers of inflammatory cells had infiltrated the mucosal epithelium at 1-month post-CBA with 30-s freezing. Therefore, we chose 7 s as the CBA treatment time in our follow-up experiments. Compared with the control group (35.81 ± 11.02 μm), BT group and CBA group (13.41 ± 4.40 μm and 4.81 ± 4.44 μm, respectively) had significantly decreased ASM thickness after 1 month (P

Nano-formulation of biosurfactant can enhance the absorption of active material. The aim of this study was to produce a nano-lipopeptide biosurfactant (NLPB) formulation from lipopeptide biosurfactant (LPB) produced by Acinetobacter junii, which was previously isolated from oil exploration areas. Besides, the wound-healing activity of the obtained NLPB formulation was also determined. The NLPB was prepared using reflux method with microwave irradiation and loaded on carbomer hydrogel formulation. The physicochemical and structural properties of NLPB formulation were evaluated using dynamic light scattering (DLS), Fourier transform infrared spectroscopy (FT-IR), and scanning electron microscopy (SEM). For determination of in vivo wound-healing activity, the prepared formulation was administered to 24 rats (previously wounded in depilated thoracic region) which were randomly distributed into six groups. The examined parameters for evaluation of the wound-healing activity were percentage wound contraction and histopathological study of regenerated tissue by hematoxylin and eosin (H&E) and trichrome staining. The size of the LPB was decreased to 80 nm by reflux method. The NLPB formulation had the most stable physicochemical properties. For the in vivo test, the concentration of 2 mg/ml was selected as the best formulation. In vivo test on rats also showed the best efficacy of the formulation on wound closure and healing. NLPB has the same physical properties as the LPB. Moreover, by decreasing the size of LPB, the wound-healing property of the LPB increased.

Background. Global longitudinal strain (GLS) can be assessed by speckle tracking echocardiography (STE) to express the degree of cardiac fibrosis. Qianyangyuyin (QYYY) granules can effectively improve GLS in hypertensive patients. Using a hypertensive rat model, we carried out speckle tracking echocardiography to validate the effect of QYYY in diminishing LV remodeling. Methods. We randomly divided 16 spontaneously hypertensive rats (SHRs) into SHR, SHR + valsartan (SHR + V), SHR + low-dose QYYY (SHR + QL), and SHR + high-dose QYYY (SHR + QH) groups, with four rats in each group. Another group of 4 Wistar-Kyoto (WKY) rats were selected into a normal control (WKY) group. At the 8th week, conventional echocardiographic parameters were measured by GE Vivid E95 ultrasound (12S probe, 10–12 MHz) and GLS by speckle tracking echocardiography with EchoPAC (version 203) software. HE and Masson’s trichrome staining were performed to detect the cardiomyocyte width and collagen volume fraction after rat sacrifice. Collagen I, α-SMA, S100A4, TGF-β, Smad 3, MYH6, and MYH7 were further analyzed by Western blot. Results. The absolute values of GLS significantly increased in the SHR + QH group compared to the SHR group, while the CVF and CW values significantly decreased. In addition, Collagen I, α-SMA, S100A4, TGF-β, Smad3, MYH7, and MYH7/MYH6 ratio remarkably reduced in the SHR + QH group. The value of GLS could be repetitively measured and positively correlated with the collagen volume fraction of the myocardium and the cardiomyocyte width of the left ventricular free wall. Conclusions. GLS is a reliable indicator to evaluate the therapeutic effect on left ventricular remodeling in hypertension. QYYY granules can inhibit the development of cardiac fibrosis in the hypertensive rat model.

Aim Peritoneal endometriosis is a chronic inflammatory disease particularly associated with macrophages. Of note, adipose tissues with fibrotic changes in the context of peritoneal endometriotic lesions are often observed during surgery. However, the characteristics of fibrotic adipose tissues in endometriosis are still unknown. In this study, we investigated the inflammatory status of retroperitoneal adipose tissues adjacent to pelvic endometriotic lesions. Methods Thirty-two patients who underwent surgical treatment were assigned to either the endometriosis (n = 16) or the control (n = 16) groups. Retroperitoneal adipose tissues around the uterus were collected from patients in both groups. Fibrosis was evaluated via Masson's trichrome staining. Macrophage infiltration, the expression of fatty acid-binding protein 4 (FABP4), and angiogenesis in the retroperitoneal adipose tissues were evaluated via immunohistochemistry. The mRNA expression levels of cytokines was also evaluated in the adipose tissues using real-time PCR. Results There was more fibrosis and angiogenesis in the adipose tissues adjacent to the endometriotic lesions with a significantly higher level of infiltration of macrophages and a predominance of the M1 type in the endometriosis group compared to the control group. In addition, FABP4 positivity in the adipose tissues of the peritoneum was significantly higher in the endometriosis group versus the control group. Moreover, the mRNA expression levels of FABP4, VEGF, and proinflammatory cytokines were also significantly higher in the endometriosis group. Conclusion Altogether, our results showed that the adipose tissue adjacent to endometriotic lesions are inflamed with fibrosis and angiogenesis.

Intestinal ischemia-reperfusion (II/R) develops when the blood flow to the intestines decreases, followed by the reestablishment of the blood supply to the ischemic tissue, resulting in intestinal mucosal barrier dysfunction, with consequent severe local and systemic inflammation. Acute lung injury (ALI) represents the most serious complication after II/R. KYP-2047 is a selective inhibitor of prolyl oligopeptidase (POP), a serine protease involved in the release of pro-angiogenic and inflammatory molecules. The aim of the present study is to assess the effects of POP-inhibition mediated by KYP-2047 treatment in the pathophysiology of ALI following II/R. An in vivo model of II/R was performed and mice were subjected to KYP-2047 treatment (intraperitoneal, 1, 2.5 and 5 mg/kg). Histological analysis, Masson's trichrome staining, immunohistochemical, immunofluorescence, biochemical and western blots analysis were performed on ileum and lung samples. KYP-2047 treatment ameliorated histological alteration in ileum and lung, reduced collagen amount and lowered inflammatory protein levels. Moreover, TGF-β1, eNOS, VEGF and CD34 positive staining has been modulated; also, a reduction in apoptosis expression was confirmed. This research revealed the strong anti-inflammatory potential of KYP-2047 associated to its modulatory role on angiogenesis and apoptosis, suggesting POP as a novel therapeutic target for ALI after II/R.

Depleted uranium (DU) used in each of civilian and military activities and contributes to health problems. Uranium is an element of heavy metals with chemical and radiological nature that have made it useful in industry and commerce, but toxic at sufficiently high doses to both humans and the environment. Also, Lead is one of the naturally occurring environmental heavy metals. This experimental study was designed to evaluate uranium and lead-induced hepatotoxicity. Material and Methods: This study was conducted in the Medical and Radiation Research Department, Research Sector, Nuclear Material Authority, Cairo, Egypt. from September to December 2020 on 30 normal healthy adult male albino rats divided into three equal groups each of 10 rats. Control group rats were kept on a normal diet and pure distilled water (Group A). The 2 experimental groups were kept on a normal diet and rats were administered with 50 mg/kg body weight of lead acetate only for 45 days, (Group B) and rats were administered with 40 mg/kg body weight of uranyl acetate only for 45 days (Group C). Animals were sacrificed and livers were removed and used to identify microscopic changes. Specimens were stained with Hematoxylin and eosin, with Masson trichrome stain for the study of fibrous tissue. Results: Mild lymphocytic infiltration, vacuolar degeneration, and mild increase of periportal fibrosis with mild depletion of glycogen significance and partial disappearance of glycogen vacuoles were reported in animals that received lead acetate contaminated water for 6 weeks. Animals that received uranyl acetate contaminated water for 6 weeks showed hepatic changes in the form of abundant lymphocytic infiltration, increased cellular polymorphism, pyknotic nuclei, and areas of cell necrosis with evident moderate periportal fibrosis and marked vacuolar degeneration associated with marked depletion of glycogen level. Conclusion: It could be concluded that exposure to uranium as compared to lead imposes a potent toxic effect on liver cells listing as glycogen depletion, cellular infiltration, and liver architecture in the form of initiation of periportal fibrosis that may progress to liver cirrhosis.

Background: Sodium valproate (SVP) is a widely prescribed treatment for epilepsy and other neurological diseases. However, liver injury is a harmful side effect related to its usage. Oxidative stress, inflammation, and fibrosis seem to play a major role in SVP-induced hepatotoxicity. Aim of the work: This work aimed, for the first time, to study the possible protective mechanisms of ginger (GN) in modulating the hepatotoxic effects of SVP in albino rats. Material and methods: A total of 24 Sprague-Dawley adult male rats (180g - 220g) were divided into equal four groups: Group1 (control): received normal saline by gavage. Group2 (GN): received ginger (200mg/kg/day). Group3 (SVP): received SVP (300mg/kg/day). Group4 (GN+SVP): received combined ginger (200mg/kg/day) and SVP (300mg/kg/day).All medications were administrated dailyfor 14 days by gavage. At the assigned time the animals were sacrificed, the blood and tissue samples were collected and processed for biochemical, histological, and immunohistochemical studies. Result: SVP treatment induced a significant increase in alanine transaminase, aspartate transaminase, and Malondialdehyde (ALT, AST, and MDA respectively), together with a significant reduction in the antioxidant enzyme Glutathione (GHS) indicated hepatic oxidative stress damage. Histological examination revealed loss of the normal hepatic architecture, extensive fibrosis identified by Masson's trichrome staining. Moreover, the SVP group showed a significant increase in tumor necrosis factor-alpha (TNF-α) and transforming growth factor-beta (TGF-β) Immunohistochemical expression. Co-treatment with GN significantly reversed the previous changes in the GN+SVP group. Conclusion: In conclusion, ginger has hepatoprotective activity due to its anti-oxidant, anti-inflammation, and anti-fibrotic potentials.

Longitudinal strain of the right ventricular (RV) free wall (RVFWLS) assessed by 2-dimensional (2D) speckle-tracking echocardiography (STE) has been recently demonstrated to correlate with the extent of RV myocardial fibrosis (MF). However, the value of 3-dimensional (3D) STE-derived strain parameters in predicting RV MF has not been investigated in patients with end-stage heart failure (HF).This study aimed to determine which RV strain parameter assessed by 2D-STE and 3D-STE was the most reliable parameter for predicting RV MF in patients with end-stage HF against histological confirmation of MF.A total of 105 consecutive patients with end-stage HF undergoing heart transplantation were enrolled in our study. The conventional RV function parameters, 2D-RVFWLS, and 3D-RVFWLS were obtained in these patients. The degree of MF was quantified by Masson trichrome staining in RV myocardial samples. The study population was divided into 3 groups according to the degree of MF on histology.Patients with severe MF had lower 3D-RVFWLS, 2D-RVFWLS, and conventional parameters of RV function compared with those with mild and moderate MF. RV MF strongly correlated with 3D-RVFWLS (r = -0.72; p 0.001), modestly with 2D-RVFWLS (r = -0.53; p 0.001), and weakly with conventional RV function parameters (r = -0.21 to -0.49; p 0.01). 3D-RVFWLS correlated best with the degree of MF (r = -0.72 vs. -0.21 to -0.53; p 0.05) compared with 2D-RVFWLS and conventional RV function parameters. 3D-RVFWLS had the highest accuracy for detecting severe MF (area under the receiver-operating characteristic curve: 0.90 vs. 0.24-0.80; p 0.05) compared with 2D-RVFWLS and conventional RV parameters. The model with 3D-RVFWLS (R3D-RVFWLS may be the most robust echocardiographic measure for predicting the extent of RV MF in patients with end-stage HF.

Francesca Rescigno, Laura Ceriotti, Marisa Meloni VitroScreen, In Vitro Innovation Center, Milan, ItalyCorrespondence: Francesca RescignoVitroScreen, In Vitro Innovation Center, via Mosè Bianchi, 103, Milan, 20149, ItalyEmail francesca.rescigno@vitroscreen.comObjective: Dermis spheroids from different donors (40 and 50 years old) were developed from primary fibroblasts to demonstrate their capacity to synthetize and organize the main dermal structural components when cultured in 3D microenvironment, forming endogenous de novo ECM according to their potential metabolic activity.Methods: Dermis spheroids were produced from primary human dermal fibroblasts at early passages in hanging drop culture system. Dermis models were characterized in terms of spheroid diameter, PICP release, collagen III and CD44 expression.Results: An increase of collagen III synthesis (101%) was found in the young donor compared to the old donor (23.5%) after seven days of culture by immunofluorescence. The progressive ECM assembly over the time and dermis maturation was showed by Masson’s trichrome staining and by immunofluorescence for collagen III and CD44; both molecules significantly accumulated in the dermal compartment from day seven to day 10 of culture with a global decrease for both spheroid models after 21 days of culture.Conclusion: Our results showed that specific culture conditions in the 3D scaffold-free microenvironment allowed the physiological and progressive ECM assembly of miniaturized dermis models reflecting phenotypic profile features of “young” and “old” native tissue from which cells were isolated with a potential application to personalized care approaches in dermatological research on aging processes and medicine.Keywords: 3D culture, dermis, scaffold-free spheroids, intrinsic aging, extracellular matrix, tissue remodeling

Monosodium glutamate (MSG) is a flavoring food additive that’s found in many foods like chips, and fast foods. The aim of this study was to demonstrate the injurious effects of MSG on the adrenals of the female albino rats and its potentiality to cross the placental barrier carrying its injurious effects to their pups. Twenty-five female albino rats weighing about (200) g were mated then separated into two groups. The control group (10 rats), and MSG-treated Group(15 rats) received 10 g/kg MSG orally, dissolved in water at a concentration of 5 g/100 ml, starting on day 7 to 20 day of gestation.A day after delivery, animals were weighed, blood samples collected then sacrificed, and the adrenals were collected and examined by the light and electron microscope. Statistically, there was a significant increase in the body weight and decrease in adrenal weight of MSG-treated mothers and their pups, as well as a decrease in the corticosterone level in MSG-treated mothers. Histologically, MSG-treated mothers and pups showed many degenerative changes, with thinning of the capsule that was confirmed by Masson trichrome stain, a disorganized structure of Zona Golmeruloza and Zona Fasciculata, and areas of shedding and hemorrhage were noticed. Shrunken nuclei and degenerated mitochondria with disrupted cristae and penetration of the lipid droplets to form “myelin-like” structures were noticed by E/M examination. The study concluded that MSG had an unwholesome effect on the structure of the adrenal tissue of both pregnant rats and their pups

Aim This study was to investigate whether bone marrow mesenchymal stem cells (BMSCs) can repair damaged endometrium in rats and its effect on endometrial receptivity. Methods A rat model of endometrial damage was established by heat injury. BMSCs were labeled with PKH26 and were transplanted into the right uterine cavity. The endometrial thickness and fertility testing were examined to assess the repair of damaged endometrium. The mass on trichrome staining was used to assess the endometrium fibrosis. The expression of integrin avβ3 and leukemia inhibitory factor (LIF) in rat endometrium was used to evaluate the endometrial receptivity. Results After transplantation of BMSCs, the distribution of PKH26 positive cells was mainly on the damaged side in the endometrial tissues. Compared to control group, the endometrial tissue structure recovered after treatment with BMSCs. BMSCs transplantation improved the fertility of endometrial injury model rats. BMSCs decreased the area of endometrial fibrosis. The expression of integrin avβ3 and LIF in endometrium was the stronger in BMSCs treatment group than control group. Conclusion BMSCs can migrate to the endometrium and repair damaged endometrium and improve endometrium receptivity.

BACKGROUND The advantages of good biocompatibility, low degradation and low antigenicity of collagen, and the osteogenic differentiation characteristics of bone mesenchymal stem cells (BMSCs) were used to promote the recovery of bone defects using partially deproteinized bone (PDPB) by bone tissue engineering (BTE). METHODS The BMSCs were identified by examining their potential for osteogenic, lipogenic, and chondrogenic differentiation. The prepared pure PDPB was ground into bone blocks 4 × 2 × 2 mm in size, which were divided into the following groups: PDPB group, PDPB + collagen group, PDPB + collagen + BMSC group, PDPB with a composite collagen nanofilm, and BMSCs injected into the tail vein. At 2, 4, 6, and 8 weeks after surgery, the effects of the implants in the different groups on bone defect repair were continuously and dynamically observed through x-ray examination, gross specimen observation, histological evaluation, and microvascularization detection. RESULTS Postoperative x-ray examination and gross specimen observation revealed that, after 4 to 8 weeks, the external contour of the graft was gradually weakened, and the transverse comparison showed that the absorption of the graft and fusion of the defect were more obvious in PDPB + collagen + BMSC group than in PDPB group and PDPB + collagen group, and the healing was better (P < 0.05). Hematoxylin and eosin staining of histological sections showed very active proliferation of trabecular hematopoietic cells in groups PDPB + collagen + BMSC and PDPB + collagen. Masson's trichrome staining for evaluation of bone defect repair showed that the mean percent area of collagen fibers was greater in PDPB + collagen + BMSC group than in the PDPB group, with degradation of the scaffold material and the completion of repair. Immunofluorescence staining showed significantly enhanced expression of the vascular marker CD31 in group C (P < 0.05). CONCLUSIONS The proposed hybrid structure of the collagen matrix and PDPB provides an ideal 3-dimensional microenvironment for patient-specific BTE and cell therapy applications. The results showed that collagen appeared to regulate MSC-mediated osteogenesis and increase the migration and invasion of BMSCs. The combination of collagen nanofilm and biological bone transplantation with BMSC transplantation enhanced the proliferation and potential of the BMSCs for bone regeneration, successfully promoting bone repair after implantation at the defect site. This method may provide a new idea for treating clinical bone defects through BTE.

Adriamycin (ADR) is an anticancer drug with important adverse effects, one of the most important of which is nephropathy. Plantago major (PM) is a widely used medicinal plant with many pharmacologic properties. The aim of the present study was to investigate the possible renoprotective effect of PM extract on ADR-induced renal fibrosis in rats. thirty-two male rats were divided into 4 groups: Control, ADR (5 mg/kg, iv), PM 600 + ADR, and PM 1200 + ADR. ADR was injected on the 7th day of the study. At the end of the 35th day of the experiment, left kidneys and serum samples were collected. The expression of collagen IV and transforming growth factor (TGF)-β were examined by immunohistochemistry. Interstitial fibrosis was estimated using Masson's trichrome staining. Serum levels of urea and creatinine were measured by commercial kits. ADR caused a significant increase in the serum level of urea compared with the control group. Interstitial fibrosis and TGF-β and collagen IV expression in the kidney tissue were also markedly increased in the ADR-treated rats. Administration of PM (600 and 1200 mg/kg) significantly improved the renal expression of these fibrotic markers and interstitial fibrosis, as well as serum levels of urea compared with the control group. Present results suggested that hydro-alcoholic extract of PM could improve kidney fibrosis induced by ADR administration.

Background: Myocardial infarction (MI) evokes an organized remodeling process characterized by the activation and transdifferentiation of quiescent cardiac fibroblasts to generate a stable collagen rich scar. Early fibroblast activation may be amenable to targeted therapy, but is challenging to identify in vivo. We aimed to non-invasively image active fibrosis by targeting the fibroblast activation protein (FAP) expressed by activated (myo)fibroblasts, using a novel positron emission tomography (PET) radioligand [68Ga]MHLL1 after acute MI. Methods: One-step chemical synthesis and manual as well as module-based radiolabeling yielded [68Ga]MHLL1. Binding characteristics were evaluated in murine and human FAP-transfected cells, and stability tested in human serum. Biodistribution in healthy animals was interrogated by dynamic PET imaging, and metabolites were measured in blood and urine. The temporal pattern of FAP expression was determined by serial PET imaging at 7 d and 21 d after coronary artery ligation in mice as percent injected dose per gram (%ID/g). PET measurements were validated by ex vivo autoradiography and immunostaining for FAP and inflammatory macrophages. Results: [68Ga]MHLL1 displayed specific uptake in murine and human FAP-positive cells (p = 0.0208). In healthy mice the tracer exhibited favorable imaging characteristics, with low blood pool retention and dominantly renal clearance. At 7 d after coronary artery ligation, [68Ga]MHLL1 uptake was elevated in the infarct relative to the non-infarcted remote myocardium (1.3 ± 0.3 vs. 1.0 ± 0.2 %ID/g, p < 0.001) which persisted to 21 d after MI (1.3 ± 0.4 vs. 1.1 ± 0.4 %ID/g, p = 0.013). Excess unlabeled compound blocked tracer accumulation in both infarct and non-infarct remote myocardium regions (p < 0.001). Autoradiography and histology confirmed the regional uptake of [68Ga]MHLL1 in the infarct and especially border zone regions, as identified by Masson trichrome collagen staining. Immunostaining further delineated persistent FAP expression at 7 d and 21 d post-MI in the border zone, consistent with tracer distribution in vivo. Conclusion: The simplified synthesis of [68Ga]MHLL1 bears promise for non-invasive characterization of fibroblast activation protein early in remodeling after MI.

BackgroundThe procoagulant state in cancer increases the thrombotic risk, and underlying cancer could affect treatment strategies and outcomes in patients with ischemic stroke. However, the histopathological characteristics of retrieved thrombi in patients with cancer have not been well studied. This study aimed to assess the histopathological difference between thrombi in patients with and without cancer.MethodsWe studied consecutive patients with acute major cerebral artery occlusion who were treated with endovascular therapy between October 2010 and December 2016 in our single-center registry. The retrieved thrombi were histopathologically investigated with hematoxylin and eosin and Masson’s trichrome staining. The organization and proportions of erythrocyte and fibrin/platelet components were studied using a lattice composed of 10×10 squares.ResultsOf the 180 patients studied, 17 (8 women, age 76.5±11.5 years) had cancer and 163 (69 women, age 74.1±11.2 years) did not. Those with cancer had a higher proportion of fibrin/platelets (56.6±27.4% vs 40.1±23.9%, p=0.008), a smaller proportion of erythrocytes (42.1±28.3% vs 57.5±25.1%, p=0.019), and higher serum D-dimer levels (5.9±8.2 vs 2.4±4.3 mg/dL, p=0.005) compared with the non-cancer cases. Receiver operating characteristic curve analysis showed the cut-off ratio of fibrin/platelet components related to cancer was 55.7% with a sensitivity of 74.8%, specificity 58.8% and area under the curve (AUC) value of 0.67 (95% CI 0.53 to 0.81), and the cut-off ratio of erythrocyte components was 44.7% with a sensitivity of 71.2%, specificity 58.9% and AUC value of 0.66 (95% CI 0.51 to 0.80).ConclusionsThromboemboli of major cerebral arteries in patients with cancer were mainly composed of fibrin/platelet-rich components.

Background: Human amniotic epithelial cells (hAECs) are seed cells used to treat acute myocardial infarction (AMI), but their mechanism remains unclear. Methods: We cultured hAECs and extracted exosomes from culture supernatants. Next, we established a stable AMI model in rats and treated them with hAECs, exosomes, or PBS. We assess cardiac function after treatment by echocardiography. Additionally, heart tissues were collected and analyzed by Masson’s trichrome staining. We conducted the tube formation and apoptosis assays to explore the potential mechanisms. Results: Cardiac function was improved, and tissue fibrosis was decreased following implantation of hAECs and their exosomes. Echocardiography showed that the EF and FS were lower in the control group than in the hAEC and exosome groups, and that the LVEDD and LVESD were higher in the control group (P

Striae distensae (SD) are linear scar tissue in the dermis that arises from stretching the skin. There are two types of SD, striae rubrae and striae albae (SA) which is marked with a line hypopigmentation that cause a psychological problem, especially in the high skin type. The standard therapy is a 0.1% tretinoin cream, but it takes a long time to reach the collagen so it needs invasive measures to reach these targets. This study was comparing 0.1% tretinoin cream with combination therapy of fractional microneedle radiofrequency (FMR) and fractional CO2 laser for SA. The number of subjects in this study was 11 patients each group with Fitzpatrick skin types IV or V. Biopsy was taken before and after therapy. The clinical changes in SA lesions in this study were represented by measurements of the length and width. The percentage of collagen area is the amount of collagen fibers with Masson's trichrome staining. The clinical changes of lesion in the combination therapy decreased. The percentage change in collagen area in the combination therapy group had a higher increase. The side effect experienced by all combination group subjects was postinflammatory hyperpigmentation, two subjects in 0.1% tretinoin cream group experienced irritation.

Objective To study the curative effect of bionic tiger‐bone powder on osteoporosis in ovariectomized rats and investigate its mechanism. Methods Overall, a 120 female Wistar rats were randomly divided into Sham (sham‐operated group), ovariectomy (OVX, ovariectomized group), TB (bionic tiger‐bone powder treatment group after ovariectomy) and TB + VD groups (bionic tiger‐bone powder + vitamin D treatment group after ovariectomy). The osteoporotic rat model was established 3 months after ovariectomy, and rats were intragastrically administrated with the corresponding drugs. Serum and bone tissue samples were collected from 10 rats in each group at weeks 4, 12 and 24 after intragastric administration. The bone microstructure of L6 vertebrae was analyzed by MicroCT, the biomechanical strength of left femurs was measured by the three‐point bending test, and serum bone metabolism markers (P1NP and CTX) were detected by ELISA. Changes in bone collagen were analyzed by Masson's trichrome staining and hydroxyproline detection, and members of the BMP2/SMAD/RUNX2 and OPG/RANKL/RANK signal pathways were detected by immunoblotting. Results Compared with the OVX group, the serum level of P1NP in the TB and TB + VD groups was higher (P, Bionic tiger‐bone powder promotes osteogenesis by activating the BMP2/SMAD/RUNX2 signaling pathway, suppresses osteoclasts by downregulating the OPG/RANK/RANKL signaling pathway, increases bone collagen content, and improves bone microstructure and bone biomechanical strength.

Peritoneal dialysis (PD) is essential for patients with end-stage renal disease. Peritoneal fibrosis (PF) is a complex inflammatory, fibrogenic process. No effective treatments are available to prevent these processes. Hepatocyte growth factor (HGF) possesses anti-inflammatory and anti-fibrotic properties. The aim of this study was to analyze whether HGF suppresses MGO-induced peritoneal inflammation and fibrosis in a mouse model. PF was induced by intraperitoneal (IP) injections of MGO for 14 days. C57/BL/6 mice were divided into three groups: Sham group (only vehicle); Sham + MGO group (PF induced by MGO); and HGF + MGO group (PF mice treated with recombinant human-HGF). PF was assessed from tissue samples by Masson’s trichrome staining. Inflammation and fibrosis-associated factors were assessed by immunohistochemistry and quantitative real-time PCR. MGO-injected mice showed significant thickening of the submesothelial compact zone with PF. Treatment with HGF significantly reduced PM thickness and suppressed the expression of collagen I and III and α-SMA. Expression of profibrotic and proinflammatory cytokines (TGF-β, TNF-α, IL-1β) was reduced by HGF treatment. The number of macrophages, and M1 and M2 macrophage-related markers, such as CD86, CD206, and CD163, was reduced in HGF + MGO mice. HGF attenuates MGO-induced PF in mice. Furthermore, HGF treatment reduces myofibroblast and macrophage infiltration, and attenuates the upregulated expression of proinflammatory and profibrotic genes in peritoneal tissues. HGF might be an effective approach to prevent the development of PF in patients undergoing PD.

This study aims to investigate the effects of melamine exposure from the weaning period (21st postnatal days in rats) on liver tissue. Female Wistar albino rats (n = 18) were divided into three groups. About 0.1-ml saline was applied to the control group by gavage for 21 days from the postnatal 21st day. The second group was taken 50-mg/kg melamine (in 0.1-ml saline) and the third group was taken 75-mg/kg melamine (in 0.1-ml saline) p.o. On the postnatal 45th day, all rats were sacrificed under anesthesia. Then, liver tissues were cut into three parts and two of them placed in neutral formalin for histopathological and flow cytometric analysis, and one of them placed in 2.5% glutaraldehyde. Histopathological analysis was performed with hematoxylin & eosin, Masson trichrome, periodic acid Schiff stained sections, and also with transmission electron microscopy. Apoptosis (Annexin V positivity) was analyzed by flow cytometry. According to histopathological analysis, hepatocyte damage, sinusoidal dilatation, and inflammatory cell infiltration significantly increased in both melamine groups compared with the control group. Apoptosis significantly increased in the 50 and 75-mg melamine groups compared with the control group. In the results of transmission electron microscopy analysis, there was abnormal chromatin distribution in the hepatocyte nuclei, loss in the cristae of the mitochondria, and organelle loss in large areas in the cytoplasm in both melamine exposure groups. As result, melamine exposure from the weaning period causes liver damage with increasing doses.

The current design requirement of cutaneous wound healing is an ideal wound dressing via biodegradable property through which fibroblasts can support, migrate and proliferate. This study aimed to evaluate the effect of the synthesized hybrid scaffold based on chitosan polymer and a natural source of calcium (Ca) and vitamin D on wound healing. To achieve this purpose, three scaffolds include in chitosan (CS), eggshell + chitosan (ES/CS) and chitosan + eggshell + vitamin D (CS/ES/Vit D) were fabricated using the freeze-drying approach. Synthesized scaffolds were characterized by field emission scanning electron microscopy (FESEM), Fourier transformed infrared (FTIR), X-ray diffraction (XRD) and MTT assay. Histological examinations of the wound healing were performed using hematoxylin–eosin (H&E) and Masson's trichrome staining methods on different days in the rat full-thickness skin wound model. Finally, the effect of the synthesized scaffold was evaluated on the TGF-β1 gene expression. The results of the scaffold characterization showed that scaffolds have the homogeneity structure. The MTT assay indicated that the cultured fibroblasts on the ES/CS and Vit D/ES/CS scaffold had viability higher than those cultured on CS. Also, histological studies demonstrated an increased effect on epithelialization and collagen production and accelerated wound healing using designed scaffolds. The TGF-β1 gene expression in all scaffolds was not significantly different between test groups and controls. In general, it can be concluded that the synthesized scaffolds have the potential to accelerate wound healing and can be used as a suitable scaffold in wound management and skin regeneration.

Colpodella spp. are free-living flagellates closely related to the apicomplexans. Human infections by Colpodella sp. have been reported. A biflagellated trophozoite and cyst stage comprise the known life cycle stages of Colpodella sp. However, the process of encystation and excystation within the life cycle is unclear. Life cycle stages initiating human infections are unknown. We performed a detailed investigation of the life cycle of Colpodella sp. (ATCC 50594) in culture using Sam-Yellowe's trichrome stains and differential interference contrast (DIC) for light microscopy and fluorescence microscopy of Congo red-stained cells and investigated ultrastructure using transmission electron microscopy (TEM). We report previously undocumented stages of Colpodella sp. Asymmetric and asynchronous division was detected inside cysts by trichrome staining and by TEM. Odd-numbered juveniles and cysts containing more than four juvenile trophozoites were identified. Live imaging of active cultures captured the excystation and egress of juvenile trophozoites and confirmed the presence of multinucleate cysts. The ultrastructure of the multinucleate cyst is reminiscent of apicomplexan schizonts. Insights gained from the life cycle stages observed in culture allowed the construction of the life cycle of Colpodella sp. Knowledge of the life cycle will aid biochemical and molecular characterization of Colpodella sp. and help identify stages in human infections.

Microbeads consisting of pullulan and dextran supplemented with hydroxyapatite have recently been developed for bone tissue engineering applications. Here, we evaluate the bone formation in two different preclinical models after injection of microbeads reconstituted with either saline buffer or autologous blood. Addition of saline solution or autologous blood to dried microbeads packaged into syringes allowed an easy injection. In the first rat bone defect model performed in the femoral condyle, microcomputed tomography performed after 30 and 60 days revealed an important mineralization process occurring around and within the core of the microbeads in both conditions. Bone volume/total volume measurements revealed no significant differences between the saline solution and the autologous blood groups. Histologically, osteoid tissue was evidenced around and in contact of the microbeads in both conditions. Using the sinus lift model performed in sheep, cone beam computed tomography revealed an important mineralization inside the sinus cavity for both groups after 3 months of implantation. Representative Masson trichrome staining images showed that bone formation occurs at the periphery and inside the microbeads in both conditions. Quantitative evaluation of the new bone formation displayed no significant differences between groups. In conclusion, reconstitution of microbeads with autologous blood did not enhance the regenerative capacity of these microbeads compared to the saline buffer group. This study is of particular interest for clinical applications in oral and maxillofacial surgery.

Background Postoperative peritoneal adhesion (PPA) is regarded as fibrous bands connecting both injured abdominal wall and organs or adjacent tissues. It is associated with T helper (Th)1 and Th2 differentiation. However, the critical role of the immunopathogenesis of adhesion formation was precisely unknown. The aim of this study was to investigate the effect of a new agent polylactic acid (PLA) nanoparticles loaded with ligustrazine, that is, ligustrazine nanoparticles (LN) on PPA and identify the potential mechanism. Methods Twenty-four Sprague–Dawley rats were randomly divided into the sham, model, LN, and sodium hyaluronate (SH) groups. The structure of LN, including entrapment efficiency (EE) and loading capacity (LC), and in vitro drug release were calculated. Adhesions were scored and the Masson's trichrome staining was used to determine the collagen deposition. The expressions of TLR4, MyD88, and NF-κB were measured by qRT-PCR, immunohistochemistry, and western blot assay. Moreover, Th1-related cytokines (IFN-γ, IL-12), Th2-related cytokines (IL-4, IL-6) in the cecum tissue and serum were conducted by ELISA. Results LN had good EE, LC, and control-release delivery characters with fairly uniform diameter and spherical morphology. It could effectively prevent adhesion formation after surgery. Besides, it could reduce collagen fibers accumulation, downregulate the expression levels of TLR4, MyD88, and NF-κB, and maintain Th1/Th2 balance. Conclusions Ligustrazine nanoparticles had effective effects on Th1/Th2 balance by regulating TLR4/MyD88/NF-κB pathway in PPA rats. It may be served as a promising therapy on postoperative adhesion formation.