The PATCH study: Prevalence of Hearing Loss During Ageing and Treatment Choices in Osteogenesis Imperfecta: A Danish Nationwide Register-Based Cohort Study.

Osteogenesis imperfecta (OI) is a group of rare hereditary collagen disorders. Hearing loss (HL) is a known complication linked to changes in the bones of the middle ear seen in OI. We aimed to determine the prevalence, age at debut, incidence, and risk of HL, surgery on bones of the middle ear, and use of hearing aids. A Danish nationwide, register-based cohort study. Data were extracted from the Danish National Patient register. Anyone with an OI diagnosis between January 1st 1977 and December 31st 2018, matched 1:5 with a reference population (Ref.Pop) on birthyear and sex, were included. 864 persons (487 women) with OI were included in the study and 4276 (2330 women) in the Ref.Pop. The sub-hazard ratio (SHR) for any HL was 4.56 [95% CI 3.64-5.71], with a prevalence of 17.0% and 4.0% in the OI cohort and Ref.Pop. Median age at debut was 42 and 58 years, respectively. The risk of otosclerosis and/or surgery was higher in the OI cohort (SHR 22.51 [95% CI 12.62-40.14]), with a median age at debut of 43 and 32 years in the OI cohort and Ref.Pop, respectively. Hearing aid use was more frequent in the OI cohort (SHR 4.16 [95% CI 3.21-5.40]) than in the Ref.Pop. The median age at debut was 45 and 60 years in the OI cohort and Ref.Pop, respectively. Persons with OI have a higher risk and prevalence of HL, hearing aids, and surgery, debuting younger, and prevalence increases with age.

Pregnancy complications and birth outcome in patients with osteogenesis imperfecta - A population-based register study.

OBJECTIVE: Patients with Osteogenesis Imperfecta (OI) have varying degrees of bone fragility and increased fracture rates. There is a paucity of data related to complications to pregnancies in patients with OI and to their offspring. With this study we aim to evaluate the risk of complications to pregnancies, delivery, and offspring in pregnancies where the mother or father have OI. DESIGN: Nationwide, register-based, cohort study. SETTING: Danish health register-based data. POPULATION: All pregnancies registered in the Danish health registers where one parent has OI and a reference population of all other pregnancies in the general population from 1997 to 2018. METHODS: Descriptive epidemiology MAIN OUTCOME MEASURES: Pregnancy and delivery complications (e.g. prevalence of pre-eclampsia, eclampsia and perinatal haemorrhage), and complications in the offspring (e.g. prevalence of low birth weight, low Apgar Score, need of CPAP or NICU, prevalence of congenital malformations (using the EUROCAT classification), incidence of osteogenesis imperfecta, prevalence of birth related fractures and hospital contacts during the first year of life) from pregnancies with parental OI. RESULTS: We identified 433 OI related pregnancies among 134 mothers with OI and 73 fathers with OI. The rates of pregnancy and delivery complications were similar between the OI cohorts and the reference population. More (31 % vs 19 %) children were delivered by caesarean section in the OI cohort than in the reference population. CONCLUSION: Pregnancies, where one parent have OI, result in live births to term with very few complications.

Histological characterization of pulp regeneration using decellularized human dental pulp and mesenchymal stem cells in a feline model.

Regenerative endodontics aims to restore pulp tissues, thus preserving the vitality of the tooth. One promising approach involves the utilization of decellularized human dental pulp (DHDP) as a scaffold repopulated with Wharton's Jelly mesenchymal stem cells (WJMSCs). This study aimed to regenerate pulp tissues using DHDP and WJMSCs following pulpectomy in mature canine teeth of a feline animal model and to investigate the histological features of the regenerated pulp. A 12-month-old male domestic shorthaired felines were used as subjects. Teeth were categorized into untreated (Group 1), pulpectomy with mineral trioxide aggregate (MTA) (Group 2), and pulpectomy with DHDP-repopulated scaffold and MTA (Group 3). The animals were sacrificed six weeks post-intervention. H&E and immunohistochemistry using anti-collagen type 1 and laminin antibodies were used to stain the tissue sections. Histological examinations presented pulp-like tissues in Group 3, with tissue components similar to the structures found in Group 1. Immunohistochemical analysis demonstrated the presence of collagen type I and laminin within the regenerated tissues. The root canals of teeth in Group 2 were devoid of pulpal tissue. DHDP with WJMSCs can potentially be used for pulp regeneration, supporting the modality for developing new clinical protocols in stem cell therapy.

Biomarkers and Biochemical Indicators to Evaluate Bone Metabolism in Preterm Neonates.

The purpose of the present study was to evaluate the concentrations of some bone turnover markers in preterm neonates with uncomplicated clinical course in the first month of life. Samples from 13 preterm neonates were collected at three different times: at birth (T0) from umbilical cord blood (UCB); and at 15 (T1) and 30 (T2) days of life from peripheral blood (PB). The concentrations of calcium (Ca), phosphate (P), total alkaline phosphatase (ALP), Collagen Type 1 Amino-terminal Propeptide (PINP), osteocalcin (OC), Collagen Type 1 Carboxyl-Terminal Telopeptide (CTX) and Leptin were assessed. A statistically significant difference for ALP concentration at birth versus T1 and T2 was found. An evident increase in the median concentrations of CTX, OC and PINP from T0 to T2 were observed. A significant difference was also found for Leptin concentration at T0 compared to T1. In preterm infants, in the absence of acute or chronic medical conditions and without risk factors for metabolic bone disease (MBD) of prematurity, there is a significant increase in bone turnover markers during the first month of life. The knowledge of the variations in these markers in the first weeks of life, integrated by the variations in the biochemical indicators of bone metabolism, could help in recognizing any conditions at risk of developing bone diseases.

Collagen 1-mediated CXCL1 secretion in tumor cells activates fibroblasts to promote radioresistance of esophageal cancer.

Esophageal squamous-cell carcinoma (ESCC) is commonly treated with radiotherapy; however, radioresistance hinders its clinical effectiveness, and the underlying mechanism remains elusive. Here, we develop patient-derived xenografts (PDXs) from 19 patients with ESCC to investigate the mechanisms driving radioresistance. Using RNA sequencing, cytokine arrays, and single-cell RNA sequencing, we reveal an enrichment of cancer-associated fibroblast (CAF)-derived collagen type 1 (Col1) and tumor-cell-derived CXCL1 in non-responsive PDXs. Col1 not only promotes radioresistance by augmenting DNA repair capacity but also induces CXCL1 secretion in tumor cells. Additionally, CXCL1 further activates CAFs via the CXCR2-STAT3 pathway, establishing a positive feedback loop. Directly interfering with tumor-cell-derived CXCL1 or inhibiting the CXCL1-CXCR2 pathway effectively restores the radiosensitivity of radioresistant xenografts in vivo. Collectively, our study provides a comprehensive understanding of the molecular mechanisms underlying radioresistance and identifies potential targets to improve the efficacy of radiotherapy for ESCC.

The Role of Bacterial, Dentinal, Salivary, and Neutrophil Degradative Activity in Caries Pathogenesis.

Until recently, it was widely accepted that bacteria participate in caries pathogenesis mainly through carbohydrate fermentation and acid production, which promote the dissolution of tooth components. Neutrophils, on the other hand, were considered white blood cells with no role in caries pathogenesis. Nevertheless, current literature suggests that both bacteria and neutrophils, among other factors, possess direct degradative activity towards both dentinal collagen type-1 and/or methacrylate resin-based restoratives and adhesives, the most common dental restoratives. Neutrophils are abundant leukocytes in the gingival sulcus, where they can readily reach adjacent tooth roots or gingival and cervical restorations and execute their degradative activity. In this review, we present the latest literature evidence for bacterial, dentinal, salivary, and neutrophil degradative action that may induce primary caries, secondary caries, and restoration failure.

Oxoglaucine Suppresses Hepatic Fibrosis by Inhibiting TGFß-Induced Smad2 Phosphorylation and ROS Generation.

Hepatic fibrosis is the first stage of liver disease, and can progress to a chronic status, such as cirrhosis or hepatocellular carcinoma. Excessive production of extracellular matrix (ECM) components plays an important role in the development of fibrosis. Mechanistically, transforming growth factor beta (TGFß)-induced phosphorylation of Smad is thought to be a key signaling pathway in the development of liver fibrosis. Although the natural isoquinoline alkaloid oxoglaucine (1,2,9,10-tetramethoxy-7H-dibenzo(de,g)quinolin-7-one) exerts numerous beneficial effects, including anti-cancer, anti-inflammatory, and anti-osteoarthritic effects in diverse cell types, the effects of oxoglaucine on liver fibrosis and fibrogenic gene expression have not been fully elucidated. The aim of this study is to evaluate the signaling pathway and antifibrotic activity of isoquinoline alkaloid oxoglaucine in TFGß-induced hepatic fibrosis in vitro. Using Hepa1c1c7 cells and primary hepatocytes, we demonstrated that oxoglaucine treatment resulted in inhibition of the expression of fibrosis markers such as collagen, fibronectin, and alpha-SMA. Subsequent experiments showed that oxoglaucine suppressed TGFß-induced phosphorylation of Smad2 and reactive oxygen species (ROS) generation, without altering cell proliferation. We further determined that the increase in Smad7 by oxoglaucine treatment is responsible for the inhibition of Smad2 phosphorylation and the anti-fibrogenic effects. These findings indicate that oxoglaucine plays a crucial role in suppression of fibrosis in hepatocytes, thereby making it a potential drug candidate for treatment of liver fibrosis.

Genetic Polymorphisms in COL1A2 gene and the Risk of Tendinopathy: Case-Control Study.

Objective To evaluate the influence of polymorphisms on genes encoding type I collagen and the genetic susceptibility of tendinopathy. Methodology Case-control study involving 242 Brazilian athletes from different sports modalities (55 cases of tendinopathy and 187 controls). The polymorphisms COL1A1 (rs1107946) and COL1A2 (rs412777, rs42524, and rs2621215) were analyzed by the TaqMan system. Odds ratio (OR) with their 95% confidence intervals (CIs) were calculated using a nonconditional logistic regression model. Results The mean age was 24.0 ± 5.6 years old and 65.3% were men. Of the 55 cases of tendinopathy, 25.4% had > 1 affected tendon, the most frequent being patellar (56.3%), rotator cuff (30.9%) and elbow or hand flexors (30.9%). Age and amount of time of sports practice were associated with a higher chance of presenting tendinopathy (5 and 8 times, respectively). The frequency of variant alleles in control and case patients, respectively, was: COL1A1 rs1107946 24.0 and 29.6%; COL1A2 rs412777 36.1 and 27.8%; rs42524 17.5 and 25.9%; and rs2621215 21.3 and 27.8%. After adjusting for confounding factors (age and years of sports practice), COL1A2 rs42524 and rs2621215 polymorphisms were associated with increased risk of tendinopathy (OR = 5.5; 95%CI = 1.2-24.6 and OR = 3.9; IC95% = 1.1-13.5, respectively). The haplotype COL1A2 CGT was associated with low risk for disease development (OR = 0.5; 95%CI = 0.3-0.9). Conclusion Age (≥ 25 years old), time of sports practice (≥ 6 years) and polymorphisms in the COL1A2 gene increased the risk of developing tendinopathy.

Single Cell Secretome Analyses of Hepatic Stellate Cells: Aiming for Single Cell Phenomics.

Activated hepatic stellate cells (HSCs) that secrete large amounts of extracellular matrix (ECM) proteins, primarily collagens, are recognized as the key pathogenic cells in liver diseases. Excessive ECM accumulation results in tissue scarring, referred to as liver fibrosis, that progresses to liver cirrhosis (liver dysfunction) and hepatocellular carcinoma. Recent studies using single cell RNA sequencing have discovered various subpopulations of HSCs with high degree of heterogeneity in quiescent, activated, as well as inactive (identified during disease regression) HSCs. However, little is known about the role of these subpopulations in ECM secretion and cell-cell communication or if they respond differently to different exogenous and endogenous factors. Moreover, how the heterogenous single cell transcriptome translates into the single cell secretome and "communicatome" (cell-cell communication) remains largely underexplored. In this chapter, we describe the method (modified enzyme-linked immunosorbent spot, ELISpot) for analyzing collagen type 1 secretion of HSCs at the single cell level, enabling a deeper understanding into the HSC secretome. In the near future, we aim to develop an integrated platform with which we can study secretome of individual cells identified by immunostaining-based fluorescence-activated cell sorting derived from healthy and diseased liver. Through the use of the VyCAP 6400-microwell chip in combination with their puncher device, we aim to perform single cell phenomics by analyzing and correlating phenotype, secretome, transcriptome, and genome of the single cells.

Genetic Polymorphisms in COL1A2 gene and the Risk of Tendinopathy: Case-Control Study / Polimorfismos genéticos no gene COL1A2 eo risco de tendinopatia: Estudo de caso-controle

Abstract Objective To evaluate the influence of polymorphisms on genes encoding type I collagen and the genetic susceptibility of tendinopathy. Methodology Case-control study involving 242 Brazilian athletes from different sports modalities (55 cases of tendinopathy and 187 controls). The polymorphisms COLIAI (rs1107946) and COLIA2 (rs412777, rs42524, and rs2621215) were analyzed by theTaqMansystem. Odds ratio(OR)withtheir 95% confidence intervals (CIs) were calculated using a nonconditional logistic regression model. Results The mean age was 24.0 ± 5.6 years old and 65.3% were men. Of the 55 cases of tendinopathy, 25.4% had > 1 affected tendon, the most frequent being patellar (56.3%), rotator cuff (30.9%) and elbow or hand flexors (30.9%). Age and amount of time of sports practice were associated with a higher chance of presenting tendinopathy (5 and 8 times, respectively). The frequency of variant alleles in control and case patients, respectively, was: COLIAI rs1107946 24.0 and 29.6%; COLIA2 rs412777 36.1 and 27.8%; rs42524 17.5 and 25.9%; and rs2621215 21.3 and 27.8%. After adjusting for confounding factors (age and years of sports practice), COLIA2 rs42524and rs2621215 polymorphisms were associated with increased risk of tendinopathy (OR = 5.5; 95% CI = 1.2-24.6 and OR = 3.9; IC95% = 1.1-13.5, respectively). The haplotype COLIA2 CGT was associated with low risk for disease development (OR = 0.5; 95%CI = 0.3-0.9). Conclusion Age (≥ 25 years old), time of sports practice (≥ 6years) and polymorphisms in the COLIA2 gene increased the risk of developing tendinopathy.

Resumo Objetivo Avaliar a influência de polimorfismos nos genes que codificam o colágeno tipo I e a suscetibilidade genética da tendinopatia. Metodologia Estudo caso-controle envolvendo 242 atletas brasileiros de diferentes modalidades esportivas (55 casos de tendinopatia e 187 controles). Os polimorfismos COL1A1 (rs1107946) e COL1A2 (rs412777, rs42524 e rs2621215) foram analisados pelo sistema TaqMan. As razões de chance (OR) com seus intervalos de confiança (IC) de 95% foram calculadas usando um modelo de regressão logística não-condicional. Resultados A média de idade foi de 24,0 ± 5,6 anos e 65,3% eram homens. Dos 55 casos de tendinopatia, 25,4% apresentaram mais de um tendão acometido, sendo os maisfrequentesopatelar(56,3%),omanguitorotador(30,9%)eodocotoveloou flexores das mãos (30,9%). A idade e o tempo de prática esportiva foram associados a uma maior chance de apresentar tendinopatia (5 e 8 vezes, respectivamente). A frequência dos alelos variantes nos controles e casos, respectivamente, foi: COL1A1 rs1107946 24,0 e 29,6%; COL1A2 rs412777 36,1 e 27,8%; rs42524 17,5 e 25,9%; e rs2621215 21,3 e 27,8%. Após ajuste pelos fatores de confundimento (idade e anos de práticas esportiva), os polimorfismos COL1A2 rs42524 e rs2621215 foram associados a um risco aumentado de tendinopatia (OR = 5,5; IC95% = 1,2-24,6 e OR = 3,9; IC95% = 1,1-13,5, respectivamente). O haplótipo COL1A2 CGT foi associado a um baixo risco para desenvolvimento da doença (OR = 0,5; IC95% = 0,3-0,9). Conclusão Aidade (> 25 anos), o tempo de prática esportiva (> 6 anos) e polimorfismos no gene COL1A2 aumentaram o risco de desenvolvimento da tendino-patia.

Genetic analysis of osteogenesis imperfecta in a large Brazilian cohort.

INTRODUCTION: Osteogenesis imperfecta (OI) is a genetically and clinically heterogeneous disorder caused by disruption of type I collagen synthesis. Previous Brazilian molecular OI studies have been restricted to case reports or small cohorts. The Brazilian OI Network (BOIN) is a multicenter study collecting clinical OI treatment data from five reference centers in three regions of Brazil. OBJECTIVE: To describe the molecular analysis of a large cohort of OI registered at BOIN. METHODS: Targeted next-generation sequencing (NGS) was performed at a centralized laboratory with the Ion Torrent platform, covering 99.6 % of the coding regions of 18 OI-associated genes. Clinical information was obtained from a clinical database. RESULTS: We included 156 subjects in the molecular analyses. Variants were detected in 121 subjects: 65 (53.7 %) in COL1A1, 42 (34.7 %) in COL1A2, 2 (1.7 %) in IFITM5, one (0.8 %) in CRTAP, three (2.5 %) in P3H1, two (1.7 %) in PPIB, four (3.3 %) FKBP10, one (0.8 %) in SERPINH1, and one (0.8 %) in TMEM38B. Ninety-one distinct variants were identified, of which 26 were novel. Of the 107 variants identified in COL1A1 and COL1A2, 24.5 % cause mild OI, while the remaining 75.5 % cause moderate, severe, or lethal OI, of which 49.3 % are glycine to serine substitutions. A single variant in FKBP10 (c.179A>C; p.Gln60Pro) was found in three unrelated and non-consanguineous participants living in the same geographic area in Northeast Brazil, suggesting a possible founder effect. CONCLUSION: Consistent with the literature, 88.4 % of the subjects had a variant in the COL1A1 and COL1A2 genes, with 10 % inherited in an autosomal recessive manner. Notably, one variant in FKBP10 with a potential founder effect requires further investigation. Data from this large cohort improves our understanding of genotype-phenotype correlations for OI in Brazil.

Reversing the imbalance in bone homeostasis via sustained release of SIRT-1 agonist to promote bone healing under osteoporotic condition.

The imbalance of bone homeostasis is the root cause of osteoporosis. However current therapeutic approaches mainly focus on either anabolic or catabolic pathways, which often fail to turn the imbalanced bone metabolism around. Herein we reported that a SIRT-1 agonist mediated molecular therapeutic strategy to reverse the imbalance in bone homeostasis by simultaneously regulating osteogenesis and osteoclastogenesis via locally sustained release of SRT2104 from mineral coated acellular matrix microparticles. Immobilization of SRT2104 on mineral coating (MAM/SRT) harnessing their electrostatic interactions resulted in sustained release of SIRT-1 agonist for over 30 days. MAM/SRT not only enhanced osteogenic differentiation and mineralization, but also attenuated the formation and function of excessive osteoclasts via integrating multiple vital upstream signals (ß-catenin, FoxOs, Runx2, NFATc1, etc.) in vitro. Osteoporosis animal model also validated that it accelerated osteoporotic bone healing and improved osseointegration of the surrounding bone. Overall, our work proposes a promising strategy to treat osteoporotic bone defects by reversing the imbalance in bone homeostasis using designated small molecule drug delivery systems.

Bronchial obstruction in osteogenesis imperfecta can be detected by forced oscillation technique.

Introduction: Respiratory insufficiency is a leading cause of death in individuals with osteogenesis imperfecta (OI). However, evaluating pulmonary function in OI presents challenges. Commonly used pulmonary function tests such as spirometry and body plethysmography are sometimes difficult to perform for OI patients, and reference intervals are not always applicable. The forced oscillation technique (FOT) is a patient-friendly method for detecting respiratory abnormalities that requires no effort from the patient. Objective: This study investigates the feasibility of FOT in the evaluation of respiratory function in the clinical management of OI patients. Methods: Twelve OI patients, comprising eight with Sillence OI I, two with OI IV, and two with OI III, underwent spirometry, body plethysmography, and FOT, both pre-and post-administration of salbutamol. Results: FOT measurements exhibited consistent trends that aligned with spirometry and body plethysmography findings. The resistance at 8 Hz decreased after the administration of salbutamol, indicating that FOT is able to detect bronchial obstruction and its alleviation by medication (p < 0.05). The resonant frequency during expiration was higher than during inspiration in nearly all patients, suggesting obstructive disease. The technique gives insight into both inspiratory and expiratory impairment of pulmonary ventilation. The main FOT parameters showed a relatively high repeatability in duplicate measurements. Conclusion: Bronchial obstruction can be detected by FOT in patients with OI during quiet breathing, making it an easily executable alternative to other lung function measurements. The technique can detect the bronchodilator effect of sympathomimetic medication. It has the potential to provide information on expiratory flow limitation, pulmonary restriction, and reduced lung compliance.

Changes in bone turnover markers in adolescents with gastroesophageal reflux disease treated with lansoprazole.

Background: Proton pump inhibitors (PPIs) have been suggested to lead to bone resorption, while the effects of PPIs on the bone mineral metabolism in children has received only limited attention in literature to date. The present study investigates whether lansoprazole alters bone turnover markers in adolescents with gastroesophageal reflux disease (GERD). Patients and methods: Included in the study were adolescents aged 16-18 with GERD and a healthy volunteers group. The GERD patient group was treated with lansoprazole 30 mg once daily for eight weeks. The serum calcium, phosphorus, magnesium, alkaline phosphatase (ALP), parathormone (PTH), 25 (OH) vitamin D, osteocalcin and urinary calcium, creatinine, deoxypyridinoline (DPD), collagen type-1 crosslinked C-telopeptide (CTX) and collagen type-1 crosslinked N-telopeptide (NTX) of both groups were studied before and after the end of the treatment. Results: A comparison of the 30 patients with GERD and the 30 volunteers revealed no significant difference in the serum calcium, phosphorus, magnesium, ALP, urinary calcium/creatinine ratio, 25 (OH) vitamin D and PTH levels measured before and after the lansoprazole treatment, while the osteocalcin, DPD, CTX and NTX values were found to be higher after treatment when compared to those at pre- treatment. Conclusions: The results of this study reveal that eight weeks of treatment with 30 mg lansoprazole daily increased the bone turnover markers of CTX, NTX, DPD and osteocalcin in adolescents aged 16-18.

The role of collagen type I/III ratio in the etiology of sacrococcygeal pilonidal.

OBJECTIVE: The purpose of this study is to investigate whether there was a difference between the midline skin and the healthy skin in the lateral by means of total amount of collagen and Type I/III ratio which was the indicator of the collagen structure. MATERIAL AND METHODS: Fifty patients with pilonidal sinus disease were enrolled. Samples were prepared from the midline skin of the sinus where the holes were located and lateral skin of the resected material. RESULTS: It was determined that the lateral line had significantly more collagen intensity and a higher collagen Type I/III ratio (p < 0.001). CONCLUSIONS: One of the reasons why hair mostly pricks into the midline in the intergluteal sulcus in pilonidal sinus disease is the fact that the amount of total collagen and collagen Type I/III ratio of the midline are lower than those of the lateral tissue. Complications are more common in cases with low Type I/III ratio and low total collagen rates.

OBJETIVO: El propósito de este estudio es investigar si existía diferencia entre la piel de la línea media y la piel sana en el lateral por medio de la cantidad total de colágeno y la relación Tipo I/III que era el indicador de la estructura del colágeno. MATERIAL Y MÉTODOS: Se inscribieron 50 pacientes con enfermedad del seno pilonidal. Se prepararon muestras de la piel de la línea media del seno donde se ubicaron los orificios y de la piel lateral del material resecado. RESULTADOS: Se determinó que la línea lateral tenía significativamente más intensidad de colágeno y una mayor relación de colágeno Tipo I/III (p < 0.001). CONCLUSIONES: Una de las razones por las que el cabello se pincha principalmente en la línea media en el surco interglúteo en la enfermedad del seno pilonidal es el hecho de que la cantidad de colágeno total y la relación de colágeno tipo I/III de la línea media son menores que las del tejido lateral. Las complicaciones son más comunes en los casos con una proporción baja de Tipo I/III y tasas bajas de colágeno total.

Modelling intramuscular drug fate in vitro with tissue-relevant biomimetic hydrogels.

Parenteral administrations are a mainstay of clinical drug delivery. Intramuscular (IM) injections deposit drug directly into skeletal muscle bellies, providing rapid systemic uptake due to the highly vascularized nature of this site. The potential to inject particulate or non-aqueous materials have also made IM injections useful for long-acting formulations. These attributes have supported a plethora of medicines being approved for IM administration. Despite these many approvals across multiple pharmaceutical categories, mechanisms that control drug release from the injection site, and thus its pharmacokinetic properties, remain poorly understood. Several pre-clinical in vivo animals have been used to model IM drug fate in patients, but these approaches have not consistently predicted clinical outcomes. This lack of a predictive in vivo model and no standardized in vitro tools have limited the options of pharmaceutical scientists to rationally design formulations for IM delivery. Here, we describe a novel, tractable in vitro model informed by dominant extracellular matrix (ECM) components present at the IM injection site. Three charge variants of green florescent protein (GFP) and the impact of three common formulation components were examined in an initial test of this in vitro model. A strongly positively charged GFP was restricted in its release from hydrogels composed of ECM components type I collagen and hyaluronic acid compared to standard and strongly negatively charged GFP. Introduction of commonly used buffers (histidine or acetate) or the non-ionic surfactant polysorbate 20 altered the release properties of these GFP variants in a manner that was dependent upon ECM element composition. In sum, this Simulator of IntraMuscular Injections, termed SIMI, demonstrated distinct release profiles of a protein biopharmaceutical surrogate that could be exploited to interrogate the impact of formulation components to expedite novel drug development and reduce current dependence on potentially non-predictive pre-clinical in vivo models.

Calprotectin and N-telopeptide of Type I Collagen (NTx) as Gingival Crevicular Fluid (GCF) Biomarker in Peri-Implantitis Patients.

INTRODUCTION: Formulation of various preventive and therapeutic strategies is possible only by a better understanding of the immune-inflammatory profile of peri-implant diseases. For understanding the changes and turnover of bone, various markers have been used in the past literature, out of which, N-telopeptide of Type I Collagen (NTx) is acknowledged to be the most reliable marker. AIMS AND OBJECTIVES: Assessment of calprotectin and NTx concentration in gingival crevicular fluid (GCF) around the implant sites in subjects suffering from peri-implantitis. MATERIALS AND METHODS: In total, 70 healthy individuals were included in the present study. These patients had opted for dental implants within the last decade. After collecting the peri-implant crevicular fluid (PICF) and GCF, various examinations were carried out. PICF samples were obtained with the help of sterile paper available in the form of strips. The enzyme-linked immunosorbent assay (ELISA) technique was used for measuring the calprotectin and NTx. All the readings were obtained in nanograms per microliter of PICF. All the results were recorded and analyzed. RESULTS: The overall mean calprotectin and NTx values were observed to be in a significantly higher range within the sites suffering from peri-implantitis when compared with healthy locations. The calprotectin values and NTx levels were positively correlated with the mean values of periodontal parameters observed clinically. CONCLUSION: Both calprotectin and NTx could be used as a biomarker signifying the presence of inflammation as well as bone resorption in patients suffering from peri-implantitis.

Exosomes Derived from Secretome Human Umbilical Vein Endothelial Cells (Exo-HUVEC) Ameliorate the Photo-Aging of Skin Fibroblast.

Purpose: This is an in-vitro experimental study to analyze the effect of Exo-HUVEC on endothelial cell (CD31), cell proliferation, matrix metalloproteinase 1 (MMP-1) and collagen type 1 on irradiated fibroblast with UVB as photo-aging model. Patients and Methods: Fibroblast cultures were divided into 5 groups, namely without UVB exposure, UVB exposure 600mJ/cm2 for 80 seconds as photo-aging model, and UVB exposure +Exo-HUVEC exposure 0.1%, 0.5% and 1%. The endothelial cell was stained with a CD31 marker, MMP-1 were examined with ELISA, cell proliferation is detected using an MTT assay; meanwhile, collagen type 1 deposition and endothelial cell were measured using flowcytometry. Results: This study found positive endothelial cell marker CD31. Significant difference was found in cell proliferation, MMP-1 and collagen type 1 level between the control group with UVB irradiation and the treatment group with Exo-HUVEC (p < 0.05). Conclusion: Exo-HUVEC significantly increases cell proliferation and collagen type 1 level, while decrease MMP-1 levels on irradiated fibroblast; therefore, Exo-HUVEC ameliorate the photo-aging of skin fibroblast.

TIMP-1-expressing breast tumor spheroids for the evaluation of drug penetration and efficacy.

Abundance of stromal cells and extracellular matrix (ECM) is observed in breast cancer, acting as a barrier for drug penetration and presenting a key issue for developing efficient therapeutics. In this study, we aimed to develop a three-dimensional (3D) multicellular tumor model comprising cancer and stromal cells that could effectively mimic the drug resistance properties of breast cancer. Three different types of spheroid models were designed by co-culturing breast cancer cells (MDA-MB-231) with three different types of stromal cells: human adipose-derived stromal cells (hASCs), human bone marrow stromal cells, or human dermal fibroblasts. Compared with other models, in the hASC co-culture model, tissue inhibitor of metalloproteinases-1 (TIMP-1) was highly expressed and the activity of matrix metalloproteinases was decreased, resulting in a higher ECM deposition on the spheroid surfaces. This spheroid model showed less drug penetration and treatment efficacy than the other models. TIMP-1 silencing in hASCs reduced ECM protein expression and increased drug penetration and vulnerability. A quantitative structure-activity relationship study using multiple linear regression drew linear relationships between the chemical properties of drugs and experimentally determined permeability values. Drugs that did not match the drug-likeness rules exhibited lower permeability in the 3D tumor model. Taken together, our findings indicate that this 3D multicellular tumor model may be used as a reliable platform for efficiently screening therapeutics agents for solid tumors.