Uso de enzimas como tratamiento dermatológico regenerador de las líneas de expresión / Use of enzymes as a regenerative dermatological treatment of expression lines / Uso de enzimas como tratamento dermatológico regenerativo de linhas de expressão

INTRODUCCIÓN: el tratamiento con enzimas es una alternativa estética mínimamente invasiva para mejorar la apariencia facial y disminuir las líneas de expresión. OBJETIVO: Determinar el uso de las enzimas hialuronidasa, colagenasa y lipasa como tratamiento enzimático dermatológico para las líneas de expresión facial. MATERIALES Y MÉTODO: estudio de campo, prospectivo, población 457 pacientes que acudieron a la consulta dermatológica entre los años 2013 y 2018 para tratamiento con enzimas. El instrumento de recolección de datos fue la hoja de registro y la fuente documental las historias clínicas. El método estadístico fue descriptivo, la información se presenta en tablas y gráficos. RESULTADOS: la edad promedio de los pacientes fue de 45,2 ± 10,1 años, 40,9% recibió 2 kits de enzimas con los 3 componentes básicos de colagenasa, hialuronidasas y lipasas. Se encontró diferencia significativa en la relación de atención entre hombres y mujeres, de 1:14, es decir las mujeres acudieron más a la consulta solicitando la colocación de este tratamiento. CONCLUSIÓN: el tratamiento con enzimas aporta beneficios al incrementar la permeabilidad dérmica, aumenta el flujo sanguíneo y el drenaje linfático, disminuye los tabiques fibrosos de la celulitis, la flacidez, adiposidades, y rejuvenece el aspecto general. Por lo que se plantea como un tratamiento efectivo para disminuir las líneas de expresión.

INTRODUCTION: enzyme treatment represents a minimally invasive aesthetic alternative to improve facial appearance and decrease expression lines. OBJECTIVE: to determine the use of the enzymes hyaluronidase, collagenase and lipase as a dermatological enzyme treatment as a regenerative treatment for expression lines. METHODS: a prospective field study was conducted of a population made up of 457 patients who attended the UNIMEL dermatological consultation between 2013 and 2018 to be treated with enzymes. The data collection instrument was the record sheet and the documentary source was the medical records. The statistical method was descriptive, the information is presented in tables and graphs. RESULTS: the average age was 45.2 ± 10.1 years of age, to whom a majority of 40.9% were applied 2 kits of enzymes with the 3 basic components of collagenase, hyaluronidases and lipases to act synergistically each other enhancing functions and revitalizing the cells of the face. A significant difference was found in the care ratio between men and women, 1:14, that is, the women attended the consultation more requesting the placement of this treatment. CONCLUSION: the use of enzymes provides great benefits to increase skin permeability, increases lymphatic drainage, reduces fibrous septa of cellulite, sagging and fat, increases blood flow and rejuvenates the general appearance. So, it is proposed as an effective treatment to reduce expression lines

INTRODUÇÃO: o tratamento enzimático é uma alternativa estética minimamente invasiva para melhorar a aparência facial e diminuir as linhas de expressão. OBJETIVO: determinar o uso das enzimas hialuronidase, colagenase e lipase como tratamento enzimático dermatológico para linhas de expressão facial. MATERIAIS E MÉTODOS: estudo de campo em perspectiva, população de 457 pacientes que compareceram à consulta dermatológica entre 2013 e 2018 para tratamento enzimático. O instrumento de coleta de dados foi a folha de registros e a fonte documental foram os registros médicos. O método estatístico foi descritivo, as informações são apresentadas em tabelas e gráficos. RESULTADOS: a idade média dos pacientes foi de 45,2 ± 10,1 anos, 40,9% receberam 2 kits de enzimas com os 3 componentes básicos de colagenase, hialuronidases e lipases. Foi encontrada diferença significativa de 1:14 na relação de atenção entre homens e mulheres, ou seja, as mulheres compareceram mais frequentemente as consultas para a colocação desse tratamento do que aos homes. CONCLUSÃO: o tratamento enzimático oferece benefícios ao aumentar a permeabilidade dérmica, aumenta o fluxo sanguíneo e a drenagem linfática, reduz os septos fibrosos da celulite, flacidez, adiposidade e rejuvenesce a aparência geral. Por isso, é proposto como um tratamento eficaz para diminuir as linhas de expressão.

How Atypical Penile Curvature Influence Clinical Outcomes in Patients with Peyronie's Disease Receiving Collagenase Clostridium Histolyticum Therapy?

Therapy for patients with burns - an integrating review

SUMMARY OBJECTIVE: to identify, through an integrative review, national studies published over the last ten years highlighting products and therapies used in burns. METHODS: integrative research with studies published in the last ten years. Including clinical studies describing the use of the already established or innovative therapies in burns and the results obtained, published in national journals in the last ten years. Excluding articles published before 2007 and those that did not present results regarding the use of products in burns. RESULTS: ten articles that met the inclusion criteria were selected. Collagenase, 1% silver sulfadiazine, and porous cellulose membrane were some of the therapies cited. CONCLUSION: the casuistry was low; however, the good results obtained with porous cellulose membrane and silver nanocrystalline dressing are highlighted, since they were used in a larger number of patients in the studies evaluated.

RESUMO OBJETIVO: Identificar, por meio de revisão integrativa, estudos nacionais publicados nos últimos dez anos que destaquem produtos e terapêuticas utilizados nas queimaduras. MÉTODOS: Pesquisa integrativa com estudos publicados nos últimos dez anos. Incluídos os estudos clínicos que descreveram a utilização de terapias já consagradas ou inovadoras em queimaduras e os resultados obtidos e publicados em periódicos nacionais nos últimos dez anos. Excluídos os artigos publicados antes de 2007 e os que não apresentaram resultados quanto ao uso de produtos nas queimaduras. RESULTADOS: Selecionados dez artigos que atenderam aos critérios de inclusão, sendo colagenase, sulfadiazina de prata 1% e membrana celulósica porosa algumas das terapias descritas. CONCLUSÕES: A casuística foi baixa, porém, ressaltam-se os bons resultados obtidos com a membrana celulósica porosa e o curativo com prata nanocristalina, em virtude de terem sido utilizados em um maior número de pacientes nos estudos avaliados.

Intra-articular Recombinant Human Growth Hormone Injection Compared with Hyaluronic Acid and Placebo for an Osteoarthritis Model of New Zealand Rabbits

PURPOSE: Up to now, there is no feasible solution for stopping or reversing the degenerative process of osteoarthritis (OA). Our study evaluated the effect of intra-articular injection of growth hormone (GH) in OA-induced rabbit knees compared to hyaluronic acid (HA) and placebo. MATERIALS AND METHODS: A total of 21 male, skeletally mature, New Zealand rabbits received an intra-articular type II collagenase injection for OA induction. Two weeks later, the rabbits were randomized into three groups based on the weekly intra-articular injection to be received: GH, HA, and saline. Injections were done for three consecutive weeks. Evaluation was done at 8 weeks after treatment, clinically using the lameness period, macroscopically using the Yoshimi score and microscopically using the Mankin score. RESULTS: The shortest period of lameness was found in the GH group (15.9±2.12 days), compared to the HA group (19.4±1.72 days) and placebo group (25.0±2.94 days). There was a statistically significant difference in macroscopic scoring between groups (p=0.001) in favor of the GH group. There was also significant difference in the microscopic score between groups (p=0.001) also in favor of the GH group. CONCLUSIONS: Intra-articular injection of GH showed better clinical, macroscopic and microscopic results as compared to HA and placebo.

Collagenase Clostridium Histolyticum Injection for Dupuytren Contracture: 2-Year Follow-up

BACKGROUND: Dupuytren disease is characterized by the development of palmar fibrous tissue that can lead to fixed flexion contracture (FFC) and contribute to functional loss of the involved digits. Our goal was to investigate rates of contracture resolution and recurrence in patients who underwent enzymatic fasciotomy for Dupuytren contracture consisting of collagenase clostridium histolyticum (CCH) injection followed by passive manipulation combined with splinting and home-based therapy. METHODS: We prospectively enrolled 34 patients (44 metacarpophalangeal [MCP] and 33 proximal interphalangeal [PIP] joints) treated by one orthopaedic hand surgeon between November 2010 and November 2014. On day 1, CCH was injected into a palpable fibrous cord of the involved fingers. The next day, the finger was passively extended to its maximal corrective position. FFC was measured for each joint before injection and immediately after manipulation. Patients were instructed to wear an extension splint at night and perform stretching exercises at home and were re-evaluated at 6 weeks, 4 months, 1 year, and 2 years. Resolution was defined as improvement of contracture to ≤ 5° of neutral. Recurrence was defined as an increase in FCC of ≥ 20° after treatment. RESULTS: Immediate contracture resolution occurred in 42 of 44 MCP joints (p < 0.001), improving from 50° to 1.5°, and in 14 of 33 PIP joints (p = 0.182), improving from 44° to 16°. Four joints had recurrence within 6 weeks. Of the 48 joints with minimum 4-month follow-up (mean, 26 months), 12 had recurrence at 2-year follow-up (MCP, 6; PIP, 6). At 2-year follow-up, MCP and PIP contractures measured 17° and 35.5°, respectively. Older age and multiple digit involvement were associated with higher recurrence rates. CONCLUSIONS: CCH offers a safe, nonoperative option to correct FCC in Dupuytren disease with greater success for MCP joints compared to PIP joints. There is a tendency of reoccurrence within 2 years of treatment. Further investigation is needed to determine optimal timing of repeat CCH injection to improve upon or extend the period of contracture resolution.

Evaluación de la actividad anticolagenasa del látex de Croton lechleri / Evaluation of the anti-collagenase activity of the latex of Croton lechleri latex

Objetivo. Determinar la capacidad inhibitoria del látex de Crotón lechleri (sangre de grado) frente a la enzima colagenasa, atenuando la formación de arrugas como consecuencia del efecto hidrolítico del colágeno por parte de la enzima. Materiales y métodos. Se ha utilizado el método in vitro de Thing para probar la actividad anticolagenasa, comparando la actividad del látex con el control positivo conformado por una solución epigalocatequina (EGCG) a concentraciones de 125, 250, 500 1000 ug/mL. Resultados. El porcentaje de inhibición de Látex de Croton lechleri a concentración de 0 ug/mL es 0; de 125 ug/mL es 5,67; de 250 ug/mL es 17,34, de 500 ug/mL es 33,41 y de 1000 ug/mL es 59,53. Conclusiones. El látex de Croton lechleri exhibió una actividad anticolagenasa significativa superando al control positivo, mostrando una IC50 de 908,02 y 1892,03 ug/mL respectivamente.

Objective. Determine the inhibitory capacity of Croton lechleri latex (Dragon's Blood) against the collagenase enzyme, reducing wrinkle formation as a consequence of the hydrolytic effect of the collagen by the enzyme. Materials and methods. Thing's in vitro method has been used to test the anti-collagenase activity, compared the activity of the latex with the positive control formed by an epigallocatechin solution (EGCG) at concentrations of 125, 250, 500 1000 ug/mL. Results. The percentage of Croton lechleri latex inhibition at a concentration of 0 ug/mL is 0, 125 ug/mL is 5.67, 250 ug/mL is 17.34, 500 ug/mL is 33.41 and 1000 ug / mL is 59.53. Conclusion. It has been concluded that Croton lechleri latex has exhibited significant anti collagenase activity exceeding the positive control, showing an IC50 of 908.02 and 1892.03 ug/mL respectively.

Matrix metalloproteinases 2 and 9 in rabbits with doxorubicin-induced cardiomyopathy / Metaloproteinases de matriz 2 e 9 em coelhos com cardiomiopatia induzida pela doxorrubicina

Some studies have shown the role played by matrix metalloproteinases and their inhibitors in doxorubicin cardiotoxicity. In this study, we sought to investigate how plasma and myocardial MMP 2 and 9 perform in rabbits with doxorubicin-induced cardiomyopathy, searching for a correlation between the activity of these collagenases and cardiac remodeling. Cardiomyopathy was induced by doxorubicin given intravenously twice a week for six consecutive weeks. Plasma MMP activity and the echocardiogram were assessed at baseline, and at 15 and 45 days after first injection of doxorubicin. The myocardial activity of these enzymes was solely evaluated in nine rabbits at 45 days, and results were compared with nine healthy controls. We only identified the full-length forms of both MMP 2 and 9 throughout the study. The plasma pro-MMP 2 reduced along the deterioration of cardiac function, while the pro-MMP 9 increased significantly at T45 as compared to baseline and T15. A negative significant correlation was found to exist between the plasma activity of pro-MMP 2 and mitral E-to-mitral septal annular early diastolic velocity ratio, which is an estimate of mean left atrial pressure and congestion. Only pro-MMP 2 was found in myocardial samples, and mean activity of such enzyme was statistically lower than that recorded for healthy controls. Although no active form was documented for either collagenase, the duration of the treatment with doxorubicin played a role in the alteration of plasma pro-forms activity. However, these changes could not be associated with most echocardiographic parameters that are supportive of cardiac remodeling.(AU)

Alguns estudos já demonstraram o papel exercido pelas metaloproteinases de matriz e seus inibidores na cardiotoxicidade promovida pela doxorrubicina. Assim, este estudo teve como objetivo investigar o comportamento das MMPs 2 e 9 plasmáticas e miocárdicas em coelhos com cardiomiopatia induzida pela doxorrubicina, buscando determinar se há correlação entre a atividade dessas colagenases e o remodelamento cardíaco. A cardiomiopatia foi induzida pela doxorrubicina aplicada por via intravenosa duas vezes por semana ao longo de seis semanas consecutivas. A atividade plasmática das MMPs e o ecocardiograma foram avaliados no momento basal e aos 15 e 45 dias após a primeira aplicação da doxorrubicina. A atividade miocárdica dessas enzimas foi quantificada em apenas nove coelhos aos 45 dias e os resultados comparados com outros nove controles saudáveis. Foram identificadas apenas as formas inativas das MMPs 2 e 9 durante todo o estudo. A pro-MMP 2 plasmática reduziu à medida que a função cardiaca se deteriorou, enquanto a pro-MMP 9 aumentou significativamente em T45 quando comparada aos momentos basal e T15. Houve correlação negativa significativa entre a atividade plasmática da pro-MMP 2 e a relação entre E mitral e a velocidade anular mitral no início da diástole, um parâmetro que permite estimar a pressão atrial esquerda média e a congestão. Apenas a pro-MMP 2 foi documentada nas amostras miocárdicas dos coelhos com cardiomiopatia e atividade media dessa enzima foi estatisticamente menor que aquela observada nos controles saudáveis. Embora a forma ativa de ambas as colagenases não tenha sido identificada, o tempo de tratamento com doxorrubicina interferiu na atividade das formas inativas plasmáticas. Contudo, essas alterações não se associaram com a maioria dos parâmetros ecocardiográficos que indicam remodelamento cardíaco.(AU)

German Cockroach Extract Induces Matrix Metalloproteinase-1 Expression, Leading to Tight Junction Disruption in Human Airway Epithelial Cells

PURPOSE: Cockroach exposure is a pivotal cause of asthma. Tight junctions are intercellular structures required for maintenance of the barrier function of the airway epithelium, which is impaired in this disease. Matrix metalloproteinases (MMPs) digest extracellular matrix components and are involved in asthma pathogenesis: MMP1 is a collagenase with a direct influence on airway obstruction in asthmatics. This study aimed to investigate the mechanism by which German cockroach extract (GCE) induces MMP1 expression and whether MMP1 release alters cellular tight junctions in human airway epithelial cells (NCI-H292). MATERIALS AND METHODS: mRNA and protein levels were determined using real-time PCR and ELISA. Tight junction proteins were detected using immunofluorescence staining. Epithelial barrier function was measured by transepithelial electrical resistance (TEER). The binding of a transcription factor to DNA molecules was determined by electrophoretic mobility shift assay, while the levels of tight junction proteins and phosphorylation were determined using Western blotting. RESULTS: GCE was shown to increase MMP1 expression, TEER, and tight junction degradation. Both an inhibitor and small interfering RNA (siRNA) of MMP1 significantly decreased GCE-induced tight junction disruption. Furthermore, transient transfection with ETS1 and SP1 siRNA, and anti-TLR2 antibody pretreatment prevented MMP1 expression and tight junction degradation. An extracellular signal-regulated kinase (ERK)/mitogen-activated protein kinase (MAPK) inhibitor also blocked MMP1 release, ETS1/SP1 DNA binding, and tight junction alteration. CONCLUSION: GCE treatment increases MMP1 expression, leading to tight junction disruption, which is transcriptionally regulated and influenced by the ERK/MAPK pathway in airway epithelial cells. These findings may contribute to developing novel therapeutic strategies for airway diseases.

Expression of collagenases (matrix metalloproteinase-1, 8, 13) and tissue inhibitor of metalloproteinase-1 of retrodiscal tissue in temporomandibular joint disorder patients

OBJECTIVES: The aim of this study was to reveal how collagenases (matrix metalloproteinase [MMP]-1, 8, 13) and tissue inhibitor of metalloproteinase 1 (TIMP-1) are expressed in immunohistochemistry of retrodiscal tissue in temporomandibular joint disorder patients. MATERIALS AND METHODS: This study was conducted on 39 patients who underwent discoplasty or discectomy. Immunohistochemical staining was undertaken and expression levels of MMP-1, 8, 13, and TIMP-1 were evaluated. The status of internal derangement of disc, osteoarthritis, and joint effusion were analyzed using magnetic resonance imaging (MRI). Disc status observed during operation was also categorized. RESULTS: The more severe disc derangement was observed on MRI, the more increased expression of MMPs and TIMP-1 appeared. Regarding MMP-13 expression, 86.7% of late-stage disc displacement patients showed grade II or III. Expression level of MMPs or TIMP was not statistically significant associated with joint effusion level. In perforation and/or adhesion groups, all patients showed grade II or III expression of MMP-13. Once perforation occurred, MMP-13 showed increased expression with statistical significance. CONCLUSION: MMP-1 and MMP-13 expression seem to be related to progression of osteoarthritis whereas MMP-8 does not seem to have a specific role with regard to temporomandibular joint disorders. TIMP-1 is considered to be partly related to internal derangement rather than osteoarthritis, but it is not significant.

A Pathophysiological Validation of Collagenase II-Induced Biochemical Osteoarthritis Animal Model in Rabbit

BACKGROUND: Current dilemma working with surgically-induced OA (osteoarthritis) model include inconsistent pathological state due to various influence from surrounding tissues. On the contrary, biochemical induction of OA using collagenase II has several advantageous points in a sense that it does not involve surgery to induce model and the extent of induced cartilage degeneration is almost uniform. However, concerns still exists because biochemical OA model induce abrupt destruction of cartilage tissues through enzymatic digestion in a short period of time, and this might accompany systemic inflammatory response, which is rather a trait of RA (rheumatoid arthritis) than being a trait of OA. METHODS: To clear the concern about the systemic inflammatory response that might be caused by abrupt destruction of cartilage tissue, OA was induced to only one leg of an animal and the other leg was examined to confirm the presence of systemic degenerative effect. RESULTS: Although the cartilage tissues were rapidly degenerated during short period of time upon biochemical induction of OA, they did not accompanied with RA-like process based on the histology data showing degeneration of articular cartilage occurred only in the collagenase-injected knee joint. Scoring evaluation data indicated that the cartilage tissues in non-induced joint remained intact. Neutrophil count transiently increase between day 8 and day 16, and there were no significant change in other complete blood count profile showing a characteristics of OA disease. CONCLUSION: These study shows that biochemically induced cartilage degeneration truly represented uniform and reliable OA state.

Collagenolytic enzymes produced by fungi: a systematic review

Abstract Specific proteases capable of degrading native triple helical or denatured collagen have been required for many years and have a large spectrum of applications. There are few complete reports that fully uncover production, characterization and purification of fungi collagenases. In this review, authors searched through four scientific on line data bases using the following keywords (collagenolytic OR collagenase) AND (fungi OR fungus OR fungal) AND (production OR synthesis OR synthesize) AND (characterization). Scientific criteria were adopted in this review to classify found articles by score (from 0 to 10). After exclusion criteria, 21 articles were selected. None obtained the maximum of 10 points defined by the methodology, which indicates a deficiency in studies dealing simultaneously with production, characterization and purification of collagenase by fungi. Among microorganisms studied the non-pathogenic fungi Penicillium aurantiogriseum and Rhizoctonia solani stood out in volumetric and specific collagenase activity. The only article found that made sequencing of a true collagenase showed 100% homology with several metalloproteinases fungi. A clear gap in literature about collagenase production by fungi was verified, which prevents further development in the area and increases the need for further studies, particularly full characterization of fungal collagenases with high specificity to collagen.

Dermoabrasión como alternativa de tratamiento en quemaduras tipo AB en pacientes pediátricos. Experiencia del Servicio de Cirugía Plástica, Unidad de Quemados del Hospital Pediátrico Fernando Bareyro / Dermoabrasion for treatment in pediatric burns

La dermoabrasión en combinación con sulfadiazina de plata y lidocaína, como tratamiento para las quemaduras AB en pacientes pediátricos, presenta ventajas en cuanto a los resultados que ofrece, ya que es un método sencillo y reproducible. Como desventaja presenta que se requieren varias sesiones de tratamiento y en algunos casos, donde se produjo profundización de la lesión, se requirió cobertura con injerto de pie

Three-dimensional (3D) printing of mouse primary hepatocytes to generate 3D hepatic structure

PURPOSE: The major problem in producing artificial livers is that primary hepatocytes cannot be cultured for many days. Recently, 3-dimensional (3D) printing technology draws attention and this technology regarded as a useful tool for current cell biology. By using the 3D bio-printing, these problems can be resolved. METHODS: To generate 3D bio-printed structures (25 mm × 25 mm), cells-alginate constructs were fabricated by 3D bio-printing system. Mouse primary hepatocytes were isolated from the livers of 6–8 weeks old mice by a 2-step collagenase method. Samples of 4 × 10⁷ hepatocytes with 80%–90% viability were printed with 3% alginate solution, and cultured with well-defined culture medium for primary hepatocytes. To confirm functional ability of hepatocytes cultured on 3D alginate scaffold, we conducted quantitative real-time polymerase chain reaction and immunofluorescence with hepatic marker genes. RESULTS: Isolated primary hepatocytes were printed with alginate. The 3D printed hepatocytes remained alive for 14 days. Gene expression levels of Albumin, HNF-4α and Foxa3 were gradually increased in the 3D structures. Immunofluorescence analysis showed that the primary hepatocytes produced hepatic-specific proteins over the same period of time. CONCLUSION: Our research indicates that 3D bio-printing technique can be used for long-term culture of primary hepatocytes. It can therefore be used for drug screening and as a potential method of producing artificial livers.

Collagenase Clostridium Histolyticum in the Treatment of Peyronie's Disease: Review of a Minimally Invasive Treatment Option

Peyronie's disease (PD) is an inflammatory disorder characterized by an abnormal collagen deposition in the tunica albuginea of the penis, leading to fibrous and non-compliant plaques that can impede normal erection. Although pharmacological treatments are available, only intralesional injection therapy and surgical reconstruction have demonstrated tangible clinical efficacy in the management of this condition. Intralesional injection of collagenase clostridium histolyticum (CCH) has come to the forefront of minimally invasive treatment of PD. In this review, the authors provide an update on the safety, efficacy, and indications for CCH. The efficacy of CCH will be assessed on the basis of improvement in the severity of penile fibrosis, curvature, and pain. Numerous well-designed clinical trials and post-approval studies involving more than 1,500 patients have consistently demonstrated the efficacy and tolerability of CCH in the treatment of PD. CCH significantly decreases penile curvature and plaque consistency, as well as improves quality of life. Post-approval studies continue to demonstrate the efficacy of CCH despite broader inclusion criteria for treatment, such as the case with acute phase disease and atypical plaque deformities (i.e., ventral plaques, hourglass narrowing). CCH continues to be the gold standard for non-surgical management of stable phase PD, in the absence of strong evidence supporting oral therapy agents and ongoing evaluation of extracorporeal shockwave therapy. However, recent studies are beginning to provide precedent for the use of CCH in the management of acute phase and atypical PD.

Histological, Physical Studies after Xenograft of Porcine Ear Cartilage

BACKGROUND: Because of the relatively similar size of organs to human and the physiological and structural similarities, the use of porcine as xenograft donors is progressing very actively. In this study, we analyzed the characteristics of porcine ear cartilage and evaluated its suitability as graft material in reconstructive and cosmetic surgery. METHODS: The auricular cartilage was harvested from two pigs, and subjected to histological examination by immunohistochemical staining. To determine the collagen content, samples were treated with collagenase and weight changes were measured. After sterilization by irradiation, the samples were grafted into rats and stained with Hematoxylin and Eosin and Masson Trichrome to observe inflammation and xenograft rejection. RESULTS: In IHC staining, extracellular matrices were mainly stained with type II collagen (20.69%), keratin sulfate (10.20%), chondroitin sulfate (2.62%), and hyaluronic acid (0.84%). After collagenase treatment, the weight decreased by 68.3%, indicating that about 70% of the porcine ear cartilage was composed of collagen. Upon xenograft of the sterilized cartilages in rats, inflammatory cells were observed for up to 2 months. However, they gradually decreased, and inflammation and reject-response were rarely observed at 5 months. CONCLUSION: The porcine ear cartilage was covered with perichondrium and cellular constituents were found to be composed of chondrocytes and chondroblasts. In addition, the extracellular matrices were mainly composed of collagen. Upon xenograft of irradiated cartilage into rats, there was no specific inflammatory reaction around the transplanted cartilage. These findings suggest that porcine ear cartilage could be a useful alternative implant material for human cosmetic surgery.

Efficacy of horse chestnut leaf extract ALH-L1005 as a matrix metalloproteinase inhibitor in ligature-induced periodontitis in canine model

Matrix metalloproteinases (MMPs) are the main proteinases associated with periodontal tissue destruction and remodeling. Therefore, inhibition of host-derived MMPs has a key role in the prevention and reduction of periodontitis progression. Horse chestnut (Aesculus hippocastanum L.) extracts have been used as treatments for inflammatory disease, traditionally. This study assessed the clinical effect as a MMP inhibitor of horse chestnut leaf extract ALH-L1005 on periodontitis. ALH-L1005 was obtained from horse chestnut leaf and its MMP inhibitory activities estimated. Periodontitis was induced in beagles assigned to 4 groups and medicated for 6 weeks: low dose test (LT; ALH-L1005, 100 mg/kg/day), high dose test (HT; ALH-L1005, 200 mg/kg/day), positive control (PC; doxycycline, 10 mg/kg/day), or negative control (NC; placebo). Before and after administration, clinical indices of the teeth and MMP quantity in gingival tissues using zymography were measured. Clinical conditions of the LT, HT, and PC groups were significantly improved after 6 weeks. In zymographic evaluations, gelatinolytic and caseinolytic activities were suppressed in LT, HT, and PC groups but not in the NC group. The results suggest that ALH-L1005 could be an effective agent for clinical prevention and treatment of periodontitis by inhibiting the gelatinase and collagenase activities, which can detach periodontal ligaments from alveolar bone.

Antioxidant and Antiaging Assays of Hibiscus sabdariffa Extract and Its Compounds

Skin aging is a complex biological process due to intrinsic and extrinsic factors. Free radical oxidative is one of extrinsic factors that induce activation of collagenase, elastase and hyaluronidase. Natural product from plants has been used as antioxidant and antiaging. This study aimed to evaluate antioxidant and antiaging properties of Hibiscus sabdariffa extract (HSE) and its compounds including myricetin, ascorbic acid, and β carotene. The phytochemical of H. sabdariffa was determined using modified Farnsworth method and presence of phenols, flavonoids and tannins were in moderate content, whereas triterpenoids and alkaloids were in low content. Total phenolic content performed using Folin-Ciocalteu method, was 23.85 µg GAE/mg. Quantitative analysis of myricetin, β-carotene, and ascorbic acid of HSE was performed with Ultra-High Performance Liquid Chromatography (UHPLC) that shows 78.23 µg/mg myricetin, 0.034 µg/mg β-carotene, whilst ascorbic acid was not detected. HSE has lower activity on DPPH (IC₅₀ = 195.73 µg/mL) compared to β-carotene, the lowest in ABTS assay (IC50 = 74.58 µg/mL) and low activity in FRAP assay (46.24 µM Fe(II)/µg) compared to myricetin, β-carotene. Antiaging was measured through inhibitory activity of collagenase, elastase, and hyaluronidase. HSE had weakest collagenase inhibitory activity (IC₅₀= 750.33 µg/mL), elastase inhibitory activity (103.83 µg/mL), hyaluronidase inhibitory activity (IC₅₀ = 619.43 µg/mL) compared to myricetin, β-carotene, and ascorbic acid. HSE contain higher myricetin compared to β-carotene. HSE has moderate antioxidants and lowest antiaging activities. Myricetin is the most active both antioxidant and antiaging activities.

Efficient Method for the Rapid Purification of Nosema ceranae Spores

Nosema ceranae is an obligate intracellular fungal parasite that causes mortality in honey bees and enhances the susceptibility of honey bees to other pathogens. Efficient purification of Nosema spores from the midgut of infected honey bees is very important because Nosema is non-culturable and only seasonably available. To achieve a higher yield of spores from honey bees, in this study, we considered that the initial release of spores from the midgut tissues was the most critical step. The use of 2 mm beads along with enzymatic treatment with collagenase and trypsin enhanced the homogenization of tissues and the yield of released spores by approximately 2.95 times compared with the use of common 3 mm beads alone. The optimal time for the enzyme treatment was determined to be 1 hr as measured by the yield and viability of the spores. A one-step filtration using a filter paper with an 8–11 µm pore size was sufficient for removing cell debris. This method may be useful to purify not only N. ceranae spores but also other Nosema spp. spores.

Primary cultures of mouse small intestinal epithelial cells using the dissociating enzyme type I collagenase and hyaluronidase

The epithelium is a highly dynamic system, which plays a crucial role in the homeostasis of the intestinal tract. However, studies on the physiological and pathophysiological functions of intestinal epithelial cells (IECs) have been hampered due to lack of normal epithelial cell models. In the present study, we established a reproducible method for primary culture of mouse IECs, which were isolated from the viable small intestinal crypts of murine fetuses (on embryonic day 19), using type I collagenase and hyaluronidase in a short span of time (≤20 min). With this method, continuously growing mouse IECs, which can be subcultured over a number of passages, were obtained. The obtained cell lines formed a tight cobblestone-like arrangement, displayed long and slender microvilli, expressed characteristic markers (cytokeratin 18 and Notch-1), and generated increasing transepithelial electrical resistance and low paracellular permeability during in vitro culture. The cells also had enzymatic activities of alkaline phosphatase and sucrase-isomaltase, and secreted various cytokines (IL-1β, IL-6, IL-8, and monocyte chemoattractant protein-1), responding to the stimulation of Escherichia coli. These results show that the primary-cultured mouse IECs obtained by the method established here had the morphological and immunological characteristics of IECs. This culture system can be a beneficial in vitro model for studies on mucosal immunology and toxicology.

How has dental pulp stem cells isolation been conducted? A scoping review

Abstract: The objective of this study was to realize a scoping review the literature in order to identify the profile of DPSCs isolation and analyze the possible risk factors that could change the native behavior of these cells. An initial search was conducted using the following MeSH terms: "(dental pulp stem cell [MeSH])"; "(dental pulp [MeSH])" AND "(stem cell [MeSH])"; "("dental pulp stem cell" [MeSH]")". The electronic search was done without date restriction up to and including April 2014, in PubMed, Scopus, Scielo and ISI Web of Knowledge databases. Studies were submitted to inclusion and exclusion criteria and 222 articles were included. Data showed that over the past 15 years many studies have been conducted using DPSCs. However this is the first systematic review regarding the isolation of stem cell, and more specifically of dental pulp stem cells. The isolation of dental pulp stem cells showed great variability, hampering the development of standard protocols to achieve in vitro dental pulp stem cells with similar characteristics. This scoping review combined, for the first time, the methodologies used for dental pulp stem isolation, highlighting the most frequently used.