Joseph Michaels

Db/db mice exhibit severe wound-healing impairments compared with other murine diabetic strains in a silicone-splinted excisional wound model

The pathophysiology of diabetic wound healing and the identification of new agents to improve clinical outcomes continue to be areas of intense research. There currently exist more than 10 different murine models of diabetes. The degree to which wound healing is impaired in these different mouse models has never been directly compared. We determined whether differences in wound impairment exist between diabetic models in order to elucidate which model would be the best to evaluate new treatment strategies. Three well‐accepted mouse models of diabetes were used in this study: db/db, Akita, and streptozocin (STZ)‐induced C57BL/6J. Using an excisional model of wound healing, we demonstrated that db/db mice exhibit severe impairments in wound healing compared with STZ and Akita mice. Excisional wounds in db/db mice show a statistically significant delay in wound closure, decreased granulation tissue formation, decreased wound bed vascularity, and markedly diminished proliferation compared with STZ, Akita, and control mice. There was no difference in the rate of epithelialization of the full‐thickness wounds between the diabetic or control mice. Our results suggest that splinted db/db mice may be the most appropriate model for studying diabetic wound‐healing interventions as they demonstrate the most significant impairment in wound healing. This study utilized a novel model of wound healing developed in our laboratory that stents wounds open using silicone splints to minimize the effects of wound contraction. As such, it was not possible to directly compare the results of this study with other studies that did not use this wound model.

Bone morphogenic protein-2 gene therapy for mandibular distraction osteogenesis

Abstract:Distraction osteogenesis (DO) requires a long consolidation period and has a low but real failure rate. Bone morphogenic proteins (BMPs) accelerate bone deposition in fractures and critical-sized bone defects, but their effects on mandibular DO are unknown. We investigated the effect of local delivery of adenovirus containing the gene for BMP-2 (Adbmp-2) on mandibular DO in a rat model. Rats (n = 54) were distracted to 3 mm over 6 days. At the start of consolidation (POD 10), Adbmp-2 or adenovirus containing the lacZgene (AdlacZ) was injected directly into the distraction zone. After 1, 2, and 4 weeks of consolidation, mandibles were evaluated for amount of bone deposition. Adbmp-2-treated specimens demonstrated an increased amount of new bone formation by radiographic, histologic, and histomorphometric analysis. This study demonstrates that local, adenovirally–mediated delivery of BMP-2 can increase bone deposition during DO, potentially shortening consolidation and enhancing DO in poorly healing mandibles, such as occurs postirradiation.

Topical vascular endothelial growth factor reverses delayed wound healing secondary to angiogenesis inhibitor administration.

The prevention of new blood vessel growth is an increasingly attractive strategy to limit tumor growth. However, it remains unclear whether anti‐angiogenesis approaches will impair wound healing, a process thought to be angiogenesis dependent. Results of previous studies differ as to whether angiogenesis inhibitors delay wound healing. We evaluated whether endostatin at tumor‐inhibiting doses delayed excisional wound closure. C57/BL6J mice were treated with endostatin or phosphate‐buffered solution 3 days prior to the creation of two full‐thickness wounds on the dorsum. Endostatin was administered daily until wound closure was complete. A third group received endostatin, but also had daily topical vascular endothelial growth factor applied locally to the wound. Wound area was measured daily and the wounds were analyzed for granulation tissue formation, epithelial gap, and wound vascularity. Endostatin‐treated mice showed a significant delay in wound healing. Granulation tissue formation and wound vascularity were significantly decreased, but reepithelialization was not effected. Topical vascular endothelial growth factor application to wounds in endostatin‐treated mice resulted in increased granulation tissue formation, increased wound vascularity, and wound closure approaching that of control mice. This study shows that the angiogenesis inhibitor endostatin delays wound healing and that topical vascular endothelial growth factor is effective in counteracting this effect.

Biologic brachytherapy: ex vivo transduction of microvascular beds for efficient, targeted gene therapy.

Background: Gene therapy for cancer holds enormous therapeutic promise, but its clinical application has been limited by the inability to achieve targeted, high-level transgene expression with limited systemic toxicity. The authors have developed a novel method for delivering genes to microvascular free flaps (commonly used during reconstructive surgery) to avoid these problems. Methods: During the finite period in which a free flap is separated from the host (ex vivo), it can be perfused with extremely high titers of genetic material through the afferent artery, resulting in efficient transduction of the tissue. Before reanastomosis, unincorporated genetic material is flushed from the flap, minimizing systemic toxicity. Results: In a rodent model using an adenoviral vector containing the lacZ reporter gene, high regional expression of β-galactosidase was achieved in all the different cells in a microvascular free flap. Moreover, no β-galactosidase staining was observed outside of the transduced flap, and viral sequence was undetectable by polymerase chain reaction analysis in other tissues. Further analysis confirmed that high-level transgene expression was precisely localized to the explanted tissue, with no collateral transduction. Conclusions: Targeting gene delivery with minimal systemic toxicity is essential for successful gene therapy. This form of “biological brachytherapy” provides a new opportunity to deliver targeted therapeutic transgenes to patients undergoing reconstructive surgery and allows microvascular free flaps to perform therapeutic and reconstructive functions.

Sonographically guided percutaneous carpal tunnel release: An anatomic and cadaveric study

Minimally invasive techniques have become the standard of care for multiple procedures. This paper demonstrates both the surgeons’ capacity to perform an accurate anatomic evaluation of the hand and forearm (n = 10) and the use of this anatomic information to accurately perform sonographically guided, percutaneous carpal tunnel release using a single-portal endoscope without direct or indirect visualization in a cadaver model (n = 6). Open dissection was then performed to confirm complete ligament transection and to evaluate the surrounding structures for injury. In all 6 cadavers, the transverse carpal ligament was transected completely without injury to any surrounding structures. With further investigation, this novel technique may offer a less invasive, office-based method for the surgical treatment of carpal tunnel syndrome that may offer patients an expedited recovery.

Using genetically modified microvascular free flaps to deliver local cancer immunotherapy with minimal systemic toxicity.

Background: Clinical use of cancer gene therapy has been prevented by the inability to deliver high levels of local transgene expression with acceptable host toxicity. The authors’ laboratory has developed an ex vivo technique to genetically modify free flaps to deliver immunotherapy locally without systemic toxicity. Methods: Superficial inferior epigastric flaps were dissected in Fischer rats, perfused with a viral vector expressing the antitumor interleukin-12 (IL-12) for 1 hour, and re-anastomosed. Beneath the flaps was a bolus of 1 × 106 beta-human chorionic gonadotropin–secreting MADB-106 tumor cells. Tumor growth was monitored using beta-human chorionic gonadotropin levels (secreted by the tumor) and size. IL-12 expression in tissue was assessed by enzyme-linked immunosorbent assay. Tumor inflammatory infiltrate was assessed using immunohistologic staining (CD8 and CD161) and enzyme-linked immunosorbent assay (interferon-&ggr;). Serum levels of liver enzymes and histologic analysis were used to assess systemic toxicity. Results: IL-12 expression was confirmed in the flap and surrounding tissue. The rate of tumor growth in the IL-12–treated group was significantly suppressed compared with the control group (p < 0.001). Liver enzyme levels remained normal, and histological evaluation of the liver, lung, and spleen revealed no evidence of inflammation in the treated group. Conclusions: Using genetically modified free flaps, the authors were able to deliver IL-12 directly into the local environment of a tumor and suppress its growth without eliciting toxic systemic effects. This technique could provide valuable adjuvant treatment after oncologic surgery for soft-tissue cancers, with the transduced flap reconstructing the defect and supplying a therapeutic agent to the resected tumor bed.

Ex vivo transduction of microvascular free flaps for localized peptide delivery.

Abstract:Gene therapy is a promising modality for the treatment of soft tissue malignancies. Our laboratory has developed a novel technique of gene transfer using microvascular free flaps that addresses many of the current barriers preventing gene therapy from achieving widespread clinical use. Our previous work has demonstrated our ability to transduce free flaps with an adenovirus encoding the reporter gene lacZ. In this current study, we show that microvascular free flaps can be transduced with an adenovirus encoding the angiogenesis inhibitor endostatin with high levels of local flap expression. These transduced free flaps were able to serve as “biologic pumps” and were able to secrete endostatin into the serum as demonstrated by enzyme-linked immunosorbent assay. This form of “biologic brachytherapy” could provide a novel approach for the continuous delivery of therapeutic genes to a localized area while avoiding many of the practical obstacles currently limiting gene therapy.

Endostatin inhibits ischemia-induced neovascularization and increases ischemic tissue loss.

The impact of inhibitors of tumor angiogenesis (endostatin, angiostatin) on the neovascularization required for the healing of transferred tissue has not been examined. We investigated the effect of endostatin on the functional neovascularization of random pattern flaps. C57BL6 mice were pretreated with endostatin beginning 3 days prior to surgery (n = 10), and daily injections continued throughout the study. Dorsal random cutaneous flaps were raised in both treatment and control (saline-treated) groups. The remaining cranial attachment was divided on day 9. Oxygen tension (PO2) was measured using a microprobe on days 1, 3, 5 and 16. Flaps were harvested and the vasculature was stained with CD31 on day 16. We found that endostatin significantly decreased flap survival. Mice that were treated with endostatin had fewer CD31+ blood vessels, worse flap perfusion at all time points, and lower oxygen tensions throughout the length of the flap. These findings have potential implications for the patients undergoing antiangiogenesis therapy who require surgical reconstruction.

Hereditary Coagulopathies: Practical Diagnosis and Management for the Plastic Surgeon

BACKGROUND Venous thromboembolism is a devastating complication representing one of the major causes of postoperative death in plastic surgery. Within the scope of plastic surgery, body-contouring procedures are often considered to carry a higher risk of venous thromboembolism. Hereditary thrombophilias comprise a group of conditions defined by a genetic predisposition to thrombosis development. Collectively, hereditary thrombophilias are present in at least 15 percent of Western populations and underlie approximately half of thromboembolic events. Although the topic of venous thromboembolism is discussed widely throughout the literature, there is little published on the diagnosis and management of hereditary thrombophilias in the plastic surgery literature. The goals of this study were to present a review of the major inherited thrombophilias, to delineate the risk of these disorders, and to recommend a practical algorithm for patient screening and management before major plastic surgery. METHODS A MEDLINE search was performed from 1965 to the present to review the literature on inherited thrombophilia disorders. RESULTS Based on the English language literature and clinical experience, the authors suggest practical guidelines for screening and management of hereditary thrombophilias. A thorough medical history and preoperative evaluation are key to reducing venous thromboembolism complications. CONCLUSIONS Hereditary thrombophilias are present in a significant number of thromboembolic events. Preoperative vigilance on the part of the plastic surgeon may help to identify patients with undiagnosed hereditary thrombophilias and thereby decrease the incidence of venous thromboembolism.

Venous thromboembolism prophylaxis in the massive weight loss patient: relative risk of bleeding.

BackgroundMeasures that can reduce the incidence of venous thromboembolism (VTE) are of great clinical importance. In addition to the use of sequential compression devices (SCDs), chemoprophylaxis with low-molecular-weight heparin (LMWH) has been recommended by the American College of Chest Physicians for major general surgery procedures. There remains inconclusive evidence to support guidelines for the plastic surgery population, and some surgeons hesitate to use anticoagulation due to concerns about bleeding in broad planes of dissection. The purpose of this study was to evaluate the risk of postoperative complications secondary to chemical thromboprophylaxis in massive weight loss patients. MethodsFive hundred forty-six surgical cases were enrolled in an institutional review board–approved prospective clinical database in the 2 years before and after routine LMWH use was initiated. Inclusion required weight loss of greater than 50 lb. Group 1 had SCDs only (n = 334), whereas group 2 had SCDs and LMWH 6 hours postoperatively (n = 212). Risk of VTE was calculated and complications of LMWH administration were analyzed. ResultsThe overall risk of deep venous thrombosis and pulmonary embolism was 0.18%. There was no statistical difference between the groups (P > 0.05). Overall risk of hematoma was 5.4%, in concordance with the literature. There was no difference in hematoma risk between the groups (4.6% before and 6.6% after LMWH; P = 0.3). The transfusion rate was 8.5% before use of LMWH (group 1) and 7.6% after (group 2; P = 0.7). ConclusionsStrategies to reduce VTE rates remain important in all areas of plastic surgery. We have demonstrated no increased risk of transfusion or hematoma and a low overall incidence of VTE after implementing a chemoprophylaxis regimen. Postoperative LMWH can provide an excellent balance between VTE prophylaxis and the risk of bleeding complications.