RESUMO
Diabetics frequently suffer from chronic, nonhealing wounds. Although bacterial colonization and/or infection are generally acknowledged to negatively impact wound healing, the precise relationship between the microbial community and impaired wound healing remains unclear. Because the host cutaneous defense response is proposed to play a key role in modulating microbial colonization, we longitudinally examined the diabetic wound microbiome in tandem with host tissue gene expression. By sequencing 16S ribosomal RNA genes, we show that a longitudinal selective shift in wound microbiota coincides with impaired healing in diabetic mice (Lepr(db/db); db/db). We demonstrate a parallel shift in longitudinal gene expression that occurs in a cluster of genes related to the immune response. Further, we establish a correlation between relative abundance of Staphylococcus spp. and the expression of cutaneous defense response genes. Our data demonstrate that integrating two types of global datasets lends a better understanding to the dynamics governing host-microbe interactions.
Assuntos
Infecções Bacterianas/fisiopatologia , Diabetes Mellitus Tipo 2/fisiopatologia , Pele/fisiopatologia , Cicatrização , Animais , Bactérias/classificação , Bactérias/genética , Bactérias/crescimento & desenvolvimento , Infecções Bacterianas/genética , Infecções Bacterianas/microbiologia , Biodiversidade , Análise por Conglomerados , Diabetes Mellitus Tipo 2/genética , Feminino , Perfilação da Expressão Gênica , Interações Hospedeiro-Patógeno , Metagenoma/genética , Camundongos , Camundongos Knockout , Dados de Sequência Molecular , Dinâmica Populacional , RNA Ribossômico 16S/genética , Receptores para Leptina/genética , Receptores para Leptina/fisiologia , Análise de Sequência de DNA , Pele/metabolismo , Pele/microbiologia , Staphylococcus/genética , Staphylococcus/fisiologia , Fatores de TempoRESUMO
Diabetic skin is known to have deficient wound healing properties, but little is known of its intrinsic biomechanical properties. We hypothesize that diabetic skin possesses inferior biomechanical properties at baseline, rendering it more prone to injury. Skin from diabetic and nondiabetic mice and humans underwent biomechanical testing. Real-time PCR was performed for genes integral to collagen synthesis and degradation. MMP-2 and MMP-9, and TIMP-1 protein levels were assessed by ELISA and zymography. Collagen I and III content was assessed using Western blot analysis. At baseline, both murine and human diabetic skin was biomechanically inferior compared to nondiabetic skin, with decreased maximum stress and decreased modulus (P < 0.001 and < 0.05, respectively). Surprisingly, the expression of genes involved in collagen synthesis were significantly up-regulated, and genes involved in collagen degradation were significantly down-regulated in murine diabetic skin (P < 0.01). In addition, MMP-2 and MMP-9/TIMP-1 protein ratios were significantly lower in murine diabetic skin (P < 0.05). Collagen I levels and I:III ratios were lower in diabetic skin (P < 0.05). These findings suggest that the predisposition of diabetics to wounds may be the result of impaired tissue integrity at baseline, and are due, in part, to a defect in the regulation of collagen protein synthesis at the post-transcriptional level.
Assuntos
Complicações do Diabetes/metabolismo , Complicações do Diabetes/patologia , Pele/metabolismo , Pele/patologia , Cicatrização/fisiologia , Animais , Fenômenos Biomecânicos , Western Blotting , Colágeno/metabolismo , Diabetes Mellitus Experimental/complicações , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Experimental/patologia , Elasticidade , Ensaio de Imunoadsorção Enzimática , Humanos , Metaloproteinase 2 da Matriz/metabolismo , Metaloproteinase 9 da Matriz/metabolismo , Camundongos , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Inibidor Tecidual de Metaloproteinase-1/metabolismoRESUMO
OBJECTIVE: Impaired diabetic wound healing is associated with abnormal stromal cell-derived factor (SDF)-1α production, decreased angiogenesis, and chronic inflammation. Lentiviral-mediated overexpression of SDF-1α can correct the impairments in angiogenesis and healing in diabetic wounds. We hypothesized that SDF-1α is a critical component of the normal wound-healing response and that inhibition of SDF-1α would further delay the wound-healing process. METHODS: dB/Db diabetic mice and Db/+ nondiabetic mice were wounded with an 8-mm punch biopsy and the wounds treated with a lentiviral vector containing either the green fluorescent protein (GFP) or SDF-1α inhibitor transgene. The inhibitor transgene is a mutant form of SDF-1α that binds, but does not activate, the CXCR4 receptor. Computerized planimetry was used to measure wound size daily. Wounds were analyzed at 3 and 7 days by histology and for production of inflammatory markers using real-time polymerase chain reaction. The effect of the SDF-1α inhibitor on cellular migration was also assessed. RESULTS: Inhibition of SDF-1α resulted in a significant decrease in the rate of diabetic wound healing, (3.8 vs 6.5 cm(2)/day in GFP-treated wounds; P = .04), and also impaired the early phase of nondiabetic wound healing. SDF-1α inhibition resulted in fewer small-caliber vessels, less granulation tissue formation, and increased proinflammatory gene expression of interleukin-6 and macrophage inflammatory protein-2 in the diabetic wounds. CONCLUSIONS: The relative level of SDF-1α in the wound plays a key role in the wound-healing response. Alterations in the wound level of SDF-1α, as seen in diabetes or by SDF-1α inhibition, impair healing by decreasing cellular migration and angiogenesis, leading to increased production of inflammatory cytokines and inflammation. Inhibition of SDF-1α further impairs diabetic wound healing.
Assuntos
Quimiocina CXCL12/biossíntese , Complicações do Diabetes/metabolismo , Cicatrização , Animais , Movimento Celular , Quimiocina CXCL12/genética , Quimiocina CXCL2/genética , Complicações do Diabetes/genética , Complicações do Diabetes/patologia , Complicações do Diabetes/fisiopatologia , Modelos Animais de Doenças , Vetores Genéticos , Tecido de Granulação/metabolismo , Tecido de Granulação/patologia , Imuno-Histoquímica , Mediadores da Inflamação/metabolismo , Interleucina-6/genética , Lentivirus/genética , Camundongos , Mutação , Neovascularização Fisiológica , Reação em Cadeia da Polimerase , RNA Mensageiro/metabolismo , Receptores CXCR4/metabolismo , Fatores de TempoRESUMO
The fetal response to small tendon injury results in regenerative or scarless healing and is characterized by a markedly diminished cellular inflammatory response, lack of fibroplasia, and restoration of normal tissue architecture. We hypothesized that an increasing fetal tendon wound size would lead to increased wound inflammation and a change from regenerative to reparative healing and scar formation. We created small or large tendon wounds in early gestation fetal sheep and used histology to assess tissue architecture, immunohistochemistry to assess the cellular inflammatory response, ovine-specific gene microarrays, and real-time reverse transcription-polymerase chain reaction to measure the gene expression in response to injury. Small tendon wounds showed a regenerative healing phenotype with orderly deposition of collagen fibers while large tendon wounds showed disorderly collagen deposition consistent with scar formation. Small tendon wounds had few inflammatory cells at 7 and 28 days after injury, whereas the large wounds showed a significant inflammatory cell infiltrate at 7 days that resolved by 28 days. At 3 days, the differential expression of genes involved in the response to injury and inflammation were seen between large and small tendon wounds. By real-time polymerase chain reaction at 7 days, large tendon wounds also had significantly increased expression of interleukin-6, interleukin-8, transforming growth factor-ß1, and transforming growth factor-ß3, compared with the small wounds. Increasing the fetal tendon wound size results in increased proinflammatory gene expression, inflammatory cell infiltration, and a change from regenerative to reparative healing. This model allows the process of regenerative healing to be examined without the confounding variable of gestational age.
Assuntos
Citocinas/genética , Doenças Fetais/genética , Regulação da Expressão Gênica no Desenvolvimento , RNA/genética , Traumatismos dos Tendões/genética , Tendões/embriologia , Cicatrização/fisiologia , Animais , Citocinas/biossíntese , Modelos Animais de Doenças , Feminino , Doenças Fetais/metabolismo , Seguimentos , Imuno-Histoquímica , Fenótipo , Reação em Cadeia da Polimerase , Gravidez , Ovinos , Traumatismos dos Tendões/embriologia , Traumatismos dos Tendões/metabolismo , Tendões/metabolismoRESUMO
BACKGROUND: Duodenal switch and single anastomosis modifications continue to gain greater interest among bariatric surgeons. Limiting factors to adoption include concerns around the nutritional management, patient compliance and follow-up, and the technical challenge of the operation. The majority of techniques offered currently use a hand-sewn duodenoileostomy. This approach is limited by the steep learning curve as well as longer operating times. OBJECTIVES: We present a video demonstrating the fully stapled technique for duodenoileostomy and ileileostomy. We offer technical pearls around the technique, specifically focused on maintaining a widely patent anastomosis, open biliopancreatic limb, safe duodenal dissection, and correct loop orientation. METHODS: Laparoscopic fully stapled duodenoileostomy for duodenal switch and single anastomosis modification. SETTING: Community hospital, single institution, 3 surgeons. CONCLUSION: Triple staple offers a reproducible and safe technique for the duodenoileostomy and specifically for construction of a Roux or loop anastomosis in duodenal switch.
Assuntos
Duodenostomia/métodos , Derivação Gástrica/métodos , Ileostomia/métodos , Laparoscopia/métodos , Obesidade Mórbida/cirurgia , Grampeamento Cirúrgico/métodos , HumanosRESUMO
INTRODUCTION: Recently, a single anastomosis duodeno-ileal bypass with sleeve gastrectomy (SADI-S) has become increasingly popular for patients with BMI > 50 as a primary or staged surgery. Staging allows surgeons to do the sleeve gastrectomy (SG) first with the conversion only happening when a failure or technical challenge is identified. PURPOSE: We present the mid-term outcomes of SADI bypass surgery after SG. METHOD: A retrospective analysis was performed on a prospective database from four institutions. Ninety-six patients were identified from 2013 to 2018. Patients were divided into two groups: one had two-stage SADI because of insufficient weight loss, the second had planned two-stage SADI because of super obesity (BMI > 50 kg/m2). Incidence of complications was divided into < 30 days and > 30 days. RESULT: Of 96 patients, 3 patients were completely lost to follow-up. The mean age was 44.8 ± 11.3 years. There were no deaths or conversion to open surgery. The postoperative early complication and late complication rate was 5.3% and 6.4% respectively. At 24 months, group 2 had higher %weight loss (WL) and change in BMI units compared to group 1 with statistically significant difference. The average WL and change in BMI for entire patient's population at 24 months after 2nd stage SADI was 20.5% and 9.4 units respectively. The remission rate for DM was 93.7% with or without the use of medication. CONCLUSION: The two-stage approach to SADI-S appears technically simpler than a single compromised operation. However, this approach needs more patients to understand its limitations.
Assuntos
Anastomose Cirúrgica , Duodeno/cirurgia , Gastrectomia , Íleo/cirurgia , Adulto , Feminino , Humanos , Hiperparatireoidismo/epidemiologia , Hipertensão/cirurgia , Tempo de Internação/estatística & dados numéricos , Lipídeos/sangue , Masculino , Pessoa de Meia-Idade , Obesidade Mórbida/cirurgia , Readmissão do Paciente/estatística & dados numéricos , Complicações Pós-Operatórias , Reoperação , Estudos Retrospectivos , Estados Unidos/epidemiologia , Vitamina K 1/sangue , Redução de Peso , Zinco/sangueRESUMO
PURPOSE: Fewer complications occur when hypospadias is repaired early in childhood. We hypothesize that the production of pro-inflammatory cytokines by fibroblasts from neonatal foreskin is decreased compared with fibroblasts from older boys. We believe that these age-related differences may explain the greater risk of complications following repair in older boys. MATERIALS AND METHODS: With IRB approval, we collected 15 samples of foreskin from boys undergoing elective circumcision. They were divided into one of three groups: a neonatal group (under 28 days), an intermediate age group (6 months-1 year), and an older age group (7-17-years-olds). Fibroblasts were cultured then incubated for 16 h with serum-free medium containing 0, 0.1, 1 or 10 ng/mL of PDGF. Supernatants were analyzed for production of IL-6 and IL-8 with quantitative ELISA. Fibroblasts had RT-PCR performed for IL-6, IL-8, IL-10, TGF-ß1, TGF-ß3 and TNF-α. RESULTS: Fibroblasts from neonatal foreskin produced significantly less IL-6 and IL-8 at baseline and following stimulation with PDGF compared to the intermediate and older age groups (P < 0.01). Real-time PCR revealed greater expression of IL-6, IL-8, TNF-α and TGF-ß1 mRNA in the older age groups (P < 0.05). CONCLUSIONS: There is a clear association between age and production of pro-inflammatory cytokines by genitourinary fibroblasts. This relationship exists at baseline and following stimulation with PDGF. The dramatic difference in levels of pro-inflammatory factors may explain the observed age-associated differences in wound scarring and stricture formation following hypospadias repair. Further clinical studies are needed, however, to validate this finding.