Photosealed Neurorrhaphy Using Autologous Tissue.

Photochemical sealing of a nerve wrap over the repair site isolates and optimizes the regenerating nerve microenvironment. To facilitate clinical adoption of the technology, we investigated photosealed autologous tissue in a rodent sciatic nerve transection and repair model. Rats underwent transection of the sciatic nerve with repair performed in three groups: standard microsurgical neurorrhaphy (SN) and photochemical sealing with a crosslinked human amnion (xHAM) or autologous vein. Functional recovery was assessed at four-week intervals using footprint analysis. Gastrocnemius muscle mass preservation, histology, and nerve histomorphometry were evaluated at 120 days. Nerves treated with a PTB-sealed autologous vein improved functional recovery at 120 days although the comparison between groups was not significantly different (SN: -58.4 +/- 10.9; XHAM: -57.9 +/- 8.7; Vein: -52.4 +/- 17.1). Good muscle mass preservation was observed in all groups, with no statistical differences between groups (SN: 69 +/- 7%; XHAM: 70 +/- 7%; Vein: 70 +/- 7%). Histomorphometry showed good axonal regeneration in all repair techniques. These results demonstrate that peripheral nerve repair using photosealed autologous veins produced regeneration at least equivalent to current gold-standard microsurgery. The use of autologous veins removes costs and foreign body concerns and would be readily available during surgery. This study illustrates a new repair method that could restore normal endoneurial homeostasis with minimal trauma following severe nerve injury.

Bone Marrow Stem Cells with Tissue-Engineered Scaffolds for Large Bone Segmental Defects: A Systematic Review.

Critical-sized bone defects (CSBDs) represent a significant clinical challenge, stimulating researchers to seek new methods for successful bone reconstruction. The aim of this systematic review is to assess whether bone marrow stem cells (BMSCs) combined with tissue-engineered scaffolds have demonstrated improved bone regeneration in the treatment of CSBD in large preclinical animal models. A search of electronic databases (PubMed, Embase, Web of Science, and Cochrane Library) focused on in vivo large animal studies identified 10 articles according to the following inclusion criteria: (1) in vivo large animal models with segmental bone defects; (2) treatment with tissue-engineered scaffolds combined with BMSCs; (3) the presence of a control group; and (4) a minimum of a histological analysis outcome. Animal research: reporting of in Vivo Experiments guidelines were used for quality assessment, and Systematic Review Center for Laboratory animal Experimentation's risk of bias tool was used to define internal validity. The results demonstrated that tissue-engineered scaffolds, either from autografts or allografts, when combined with BMSCs provide improved bone mineralization and bone formation, including a critical role in the remodeling phase of bone healing. BMSC-seeded scaffolds showed improved biomechanical properties and microarchitecture properties of the regenerated bone when compared with untreated and scaffold-alone groups. This review highlights the efficacy of tissue engineering strategies for the repair of extensive bone defects in preclinical large-animal models. In particular, the use of mesenchymal stem cells, combined with bioscaffolds, seems to be a successful method in comparison to cell-free scaffolds.

Light-Activated Vascular Anastomosis.

Background. There have been few advances in technique since vascular anastomosis was performed with silk suture on a curved needle in 1902. This technique results in disruption of the endothelium with exposed intraluminal suture, both of which may lead to thrombocyte aggregation, intimal hyperplasia, and vascular stenosis. A variety of alternative techniques have been explored, with limited success. Photochemical tissue bonding (PTB) is a light-activated methodology of rapidly cross-linking tissue interfaces at the molecular level. Herein, we describe a new technique for anastomosis of venous interposition graft in an ovine model of femoral artery bypass utilizing PTB. Methods. Polypay specific pathogen free sheep (n = 5; 40-45 kg) underwent femoral artery bypass utilizing saphenous vein. The femoral artery was transected and reversed saphenous vein was implanted as an interposition graft. The proximal anastomosis was created as a vein-over-artery cuff utilizing PTB, and the distal anastomosis was created with standard interrupted 8-0 polypropylene suture. Four weeks post-index operation, femoral angiogram was performed to evaluate patency, tortuosity, and luminal diameter. All bypass grafts were harvested and longitudinal and transverse histological sections from the proximal anastomosis were analyzed. Results. The PTB anastomoses (n = 5) were immediately watertight and patent. All animals survived the 28-day study duration. Angiography revealed patent grafts with no aneurysm or stenosis (n = 5). Histologic examination revealed integration of the venous endothelium with the arterial adventitia. Conclusion. Photochemical tissue bonding creates an immediate strong, watertight vascular anastomosis that can withstand physiologic arterial pressure and remains patent at 28 days without the need for intraluminal suture.

Surgical Approaches for Prevention of Neuroma at Time of Peripheral Nerve Injury.

Painful neuroma is a frequent sequela of peripheral nerve injury which can result in pain and decreased quality of life for the patient, often necessitating surgical intervention. End neuromas are benign neural tumors that commonly form after nerve transection, when axons from the proximal nerve stump regenerate in a disorganized manner in an attempt to recreate nerve continuity. Inflammation and collagen remodeling leads to a bulbous end neuroma which can become symptomatic and result in decreased quality of life. This review covers surgical prophylaxis of end neuroma formation at time of injury, rather than treatment of existing neuroma and prevention of recurrence. The current accepted methods to prevent end neuroma formation at time of injury include different mechanisms to inhibit the regenerative response or provide a conduit for organized regrowth, with mixed results. Approaches include proximal nerve stump capping, nerve implantation into bone, muscle and vein, various pharmacologic methods to inhibit axonal growth, and mechanisms to guide axonal growth after injury. This article reviews historical treatments that aimed to prevent end neuroma formation as well as current and experimental treatments, and seeks to provide a concise, comprehensive resource for current and future therapies aimed at preventing neuroma formation.

A Photosealed Cap Prevents Disorganized Axonal Regeneration and Neuroma following Nerve Transection in Rats.

Neuroma is a common sequela of traumatic peripheral nerve injury that can result in pain and decreased quality of life for patients. Neuromas result from axonal outgrowth in an attempt to reestablish continuity with the disrupted distal nerve end. Photosealing is a light-activated technique whereby tissues can be securely isolated in a strong and secure manner. This study investigated whether photosealing of autologous vein and crosslinked human amniotic membrane (xHAM) to cap the proximal stump of transected sciatic nerve would prevent disorganized axonal regeneration and neuroma in a rat model. Methods: The right sciatic nerve of Lewis rats (n = 27, 300-350 g) was transected 1 cm proximal to the trifurcation. Animals were randomized to one of three groups (n = 9): no further intervention (Group 1), photosealing with xHAM (Group 2), or photosealing with vein (Group 3). After 60 days, rats were euthanized and their right hindlimbs were re-explored for evidence of disorganized axonal regeneration and/or bulbous neuroma. Results: All untreated control animals were found to have protruding nerve fibers, often invading the adjacent muscle, and 33% of these control animals exhibited a bulbous neuroma. Photosealing with xHAM successfully capped 100% of nerves, with no observable axonal outgrowth. Photosealing with vein prevented axonal outgrowth in eight of nine nerves. No bulbous neuroma was found in any photosealed nerves. Conclusion: Nerve capping with photosealed xHAM or autologous vein can prevent axonal outgrowth in transected nerves, therefore decreasing the likelihood of symptomatic neuroma formation following nerve transection injury or surgical intervention.

Light-activated photosealing with human amniotic membrane strengthens bowel anastomosis in a hypotensive, trauma-relevant swine model.

BACKGROUND: Gastrointestinal anastomotic leakage is a dreaded complication despite advancements in surgical technique. Photochemical tissue bonding (PTB) is a method of sealing tissue surfaces utilizing photoactive dye. We evaluated if crosslinked human amniotic membrane (xHAM) photosealed over the enteroenterostomy would augment anastomotic strength in a trauma-relevant swine hemorrhagic shock model. METHODS: Yorkshire swine (40-45 kg, n = 14) underwent midline laparotomy and sharp transection of the small intestine 120 cm proximal to the ileocecal fold. Immediately following intestinal transection, a controlled arterial bleed was performed to reach hemorrhagic shock. Intestinal repair was performed after 60 minutes and autotransfusion of the withdrawn blood was performed for resuscitation. Animals were randomized to small intestinal anastomosis by one of the following methods (seven per group): suture repair (SR), or SR with PTB augmentation. Animals were euthanized at postoperative Day 28 and burst pressure (BP) strength testing was performed on all excised specimens. RESULTS: Mean BP for SR, PTB, and native tissue groups were 229 ± 40, 282 ± 21, and 282 ± 47 mmHg, respectively, with the SR group statistically significantly different on analysis of variance (p = 0.02). Post-hoc Tukey all-pairs comparison demonstrated a statistically significant difference in burst pressure strength between the SR only and the PTB group (p = 0.04). All specimens in SR group ruptured at the anastomosis upon burst pressure testing, while all specimens in the PTB group ruptured at least 2.5 cm from the anastomosis. CONCLUSION: Photosealing with xHAM significantly augments the strength of small intestinal anastomosis performed in a trauma porcine model.

Photochemical Tissue Passivation of Arteriovenous Grafts Prevents Long-Term Development of Intimal Hyperplasia in a Swine Model.

BACKGROUND: The autologous vein remains the standard conduit for lower extremity and coronary artery bypass grafting despite a 30%-50% 5-y failure rate, primarily attributable to intimal hyperplasia (IH) that develops in the midterm period (3-24 mo) of graft maturation. Our group discovered that externally strengthening vein grafts by cross-linking the adventitial collagen with photochemical tissue passivation (PTP) mitigates IH in an arteriovenous model at 4 wk. We now investigate whether this effect is retained in the midterm period follow-up. METHODS: Six Hanford miniature pigs received bilateral carotid artery interposition vein grafts. In each animal, the external surface of one graft was treated with PTP before grafting, whereas the opposite side served as the untreated control. The grafts were harvested after 3 mo. Ultrasound evaluation of all vein grafts was performed at the time of grafting and harvest. The grafts were also evaluated histomorphometrically and immunohistologically for markers of IH. RESULTS: All vein grafts were patent at 3 mo except one graft in the PTP-treated group because of early technical failure. The control vein grafts had significantly greater IH than PTP-treated grafts at 3 mo, as evidenced by the intimal area (2.6 ± 1.0 mm2versus 1.4 ± 1.5 mm2, respectively, P = 0.045) and medial area (5.1 ± 1.9 mm2versus 2.7 ± 2.4 mm2, respectively, P = 0.048). The control grafts had an increased presence and proliferation of mural myofibroblasts with greater smooth muscle actin and proliferating cell nuclear antigen staining. CONCLUSIONS: PTP treatment to the external surface of the vein grafts decreases IH at 3 mo after arteriovenous grafting and may prevent future graft failure.

Photochemical Tissue Passivation Prevents Contracture of Full Thickness Wounds in Mice.

BACKGROUND AND OBJECTIVES: Wound contracture formation from excessive myofibroblast activity can result in debilitating morbidities. There are currently no treatments to prevent contracture. Photochemical tissue passivation (PTP), an established, safe, and user-friendly treatment modality, crosslinks collagen by a light-activated process, thus modulating the wound healing response and scarring. We hypothesised that PTP treatment would reinforce wounds by blunting the fibrotic response thus limiting contracture. STUDY DESIGN/MATERIALS AND METHODS: Full-thickness, 1 cm × 1 cm excisional wounds were created on the dorsum of 32 C57BL/6 mice. Treated wounds were painted with photosensitizing dye and exposed to visible light. Wounds were serially photographed over 6 weeks to measure wound contracture. At 7, 14, 21, and 42 days after wound creation, mice were euthanized and wounds were harvested for histologic review by a dermatopathologist. RESULTS: By Day 7, control wounds had significantly more contracture than those treated with PTP (33.0 ± 17.1% and 19.3 ± 9.0%, respectively; P = 0.011). PTP-treated wounds maintained approximately 20% less contracture than controls from Day 14 and on (P < 0.05). By Day 42, wounds had contracted by 86.9 ± 5.5% in controls and 64.2 ± 3.2% in PTP-treated wounds (P < 0.03). Histologically, PTP wounds had earlier growth and development of dermal collagen, neovascularization, and development of skin appendages, compared with control wounds. CONCLUSIONS: PTP significantly limits contracture of full-thickness wounds and improves wound healing. PTP-treated wounds histologically demonstrate more mature structural organization than untreated wounds and closely resemble native skin. PTP treatment may be applicable not only for excisional wounds, but also for wounds with a high incidence of contracture and associated morbidity. Lasers Surg. Med. © 2019 Wiley Periodicals, Inc.

Medical Applications of Rose Bengal- and Riboflavin-Photosensitized Protein Crosslinking.

This review summarizes research on many of the potential applications of photosensitized crosslinking of tissue proteins in surgery and current knowledge of the photochemical mechanisms underlying formation of the covalent protein-protein crosslinks involved. Initially developed to close wounds or reattach tissues, protein photocrosslinking has also been demonstrated to stiffen and strengthen tissues, decrease inflammatory responses and facilitate tissue bioengineering. These treatments appear to result largely from crosslinks within and between collagen molecules in tissue that typically form by an oxygen-dependent mechanism. Surgical applications discussed include sealing wounds in skin, cornea and bowel; reattaching severed nerves, blood vessels and tendons; strengthening cornea and vein; reducing capsular contracture after breast implants; and regenerating joint cartilage.

Photochemical Tissue Passivation Attenuates AV Fistula Intimal Hyperplasia.

OBJECTIVE: We hypothesized that decreasing vein compliance would protect the vein against stretch injury and reduce intimal hyperplasia (IH). BACKGROUND: Although arteriovenous fistulas (AVFs) are the criterion standard for vascular access, their effectiveness is limited by poor patency with 40% to 60% failing due to IH. Venous stretch injury from exposure to arterial pressure induces IH. Photochemical tissue passivation (PTP) crosslinks adventitial collagen, decreasing vein compliance to resemble that of an artery. METHODS: AVFs were created between the femoral artery and epigastric vein in rats (n = 29). PTP was performed on the vein immediately before vessel anastomosis. AVFs were harvested after four weeks. Venous diameter was measured at the initial procedure and harvest. Intimal area was measured for each segment. Ultrasound was performed at harvest to measure AVF flow. RESULTS: Following AVF construction, venous diameter increased by 10% ±â€Š18% for PTP-treated vessels and 78% ±â€Š27% for controls (P ≤ 0.0001). At one month, PTP reduced AVF dilation by 71% compared to control (69% ±â€Š29% vs 241% ±â€Š78%; P ≤ 0.0001). Both juxta-anastomotic intimal area and total intimal area were reduced in PTP-treated vessels compared to control vessels. Specifically, intimal area was 0.024 ±â€Š0.018 and 0.095 ±â€Š0.089 mm for PTP-treated juxta-anastomotic segments of AVF and control, respectively (P < 0.05). Mean total intimal area for PTP-treated and control AVF were 0.080 ±â€Š0.042 and 0.190 ±â€Š0.110 mm, respectively (P < 0.03). AVF flow was 46.9 ±â€Š35.3 and 19.1 ±â€Š10.1 mL/min for PTP-treated and control AVF, respectively (P < 0.109). CONCLUSIONS: These data demonstrate that PTP represents a promising therapy for the prevention of AVF IH, a process that might improve surgical outcomes for patients receiving hemodialysis.

An intraluminal stent facilitates light-activated vascular anastomosis.

BACKGROUND: Photochemical tissue bonding (PTB) is a sutureless, light-activated technique that produces a watertight, microvascular repair with minimal inflammation compared to standard microsurgery. However, it is practically limited by the need for a clinically viable luminal support system. The aim of this study was to evaluate a hollow biocompatible stent to provide adequate luminal support to facilitate vascular anastomosis using the PTB technique. METHODS: Forty rats underwent unilateral femoral artery transection. Five rats were used to optimize the stent delivery method, and the remaining 35 rats were randomized into three groups: (1) standard suture repair with 10-0 nylon microsuture (SR), (2) standard suture repair over the stent (SR + S), or (3) PTB repair over stent (PTB + S). For the PTB group, a 2-mm overlapping cuff was painted with 0.1% (wt/vol) Rose Bengal then illuminated for 30 seconds on each side (532 nm, 0.5 W/cm, 30 J/cm). Anastomotic leak and vessel patency (immediate, 1 hour, and 1 week postoperatively) were assessed. RESULTS: Vessels in all three groups were patent immediately and at 1 hour postoperatively. After 1 week, all animals displayed patency in the SR group, while only 5 of 14 and 2 of 8 surviving animals had patent vessels in the PTB + S and SR + S groups, respectively. CONCLUSIONS: This study demonstrated successful use of an intraluminal stent for acute microvascular anastomosis using the PTB technique. However, the longer-term presence of the stent at the anastomotic site led to thrombosis in multiple cases. A rapidly dissolvable stent should facilitate a light-activated microvascular anastomosis with excellent long-term patency.

Prevention of vein graft intimal hyperplasia with photochemical tissue passivation.

OBJECTIVE: Saphenous vein is the conduit of choice for bypass grafting. Saphenous vein grafts have poor long-term patency rates because of intimal hyperplasia (IH) and subsequent accelerated atherosclerosis. One of the primary triggers of IH is endothelial injury resulting from excessive dilation of the vein after exposure to arterial pressures. Photochemical tissue passivation (PTP) is a technology that cross-links adventitial collagen by a light-activated process, which limits dilation by improving vessel compliance. The objective of this study was to investigate whether PTP limits the development of IH in a rodent venous interposition graft model. METHODS: PTP is accomplished by coating venous adventitia with a photosensitizing dye and exposing it to light. To assess the degree of collagen cross-linking after PTP treatment, a biodegradation assay was performed. Venous interposition grafts were placed in the femoral artery of Sprague-Dawley rats. Rats were euthanized after 4 weeks, and intimal thickness was measured histologically. Vein dilation at the time of the initial procedure was also measured. RESULTS: Time to digestion was 63 ± 7 minutes for controls, 101 ± 2.4 minutes for rose bengal (RB), and 300 ± 0 minutes for PTP (P < .001 PTP vs control). A total of 37 animals underwent the procedure: 12 PTP, 12 RB only, and 13 untreated controls. Dilation of the graft after clamp release was 99% for control, 65% for RB only, and 19% for PTP-treated (P < .001 PTP vs control). Intimal thickness was 77 ± 59 µm in controls, 60 ± 27 µm in RB only, and 33 ± 28 µm in PTP-treated grafts. There was a statistically significant 57% reduction in intimal thickness after treatment with PTP compared with untreated controls (P = .03). CONCLUSIONS: PTP treatment of venous interposition grafts in a rat model resulted in significant collagen cross-linking, decreased vessel compliance, and significant reduction in IH.

No midterm advantages in the middle term using small intestinal submucosa and human amniotic membrane in Achilles tendon transverse tenotomy.

BACKGROUND: The study was aimed to compare the effects of small intestinal submucosa (SIS) and human amniotic membrane (HAM) on Achilles tendon healing. METHODS: A total of 48 New Zealand white rabbits were divided into two groups. A full-thickness transverse tenotomy was made at the right leg of the rabbits. Then, the laceration site was wrapped with HAM (P/A group) or SIS (P/S group). The ultimate stress (US) and Young's modulus (E) of the tendons were detected for biomechanical analysis. Histological evaluation was performed using hematoxylin and eosin, immunohistochemical, and immunofluorescent stain. Expression of collagen I was detected by western blot analysis, and levels of inflammatory cytokines IL-1ß, IL-6, and TNF-α were measured. Finally, adhesion formation was evaluated. RESULTS: There were no significant differences in filamentous adhesion, cross-sectional areas of the laceration sites, levels of inflammatory response, and collagen type I expression between the P/A and P/S groups (p > 0.05). Compared with the P/A group, the US and E values were significantly higher in the P/S group at day 7 (p < 0.05) and at day 14 (p < 0.05). In addition, vascularity was significantly higher in the P/S group than that in the P/A group at day 3 (p < 0.05), day 7 (p < 0.01), and day 9 (p < 0.05). CONCLUSIONS: SIS showed superior biomechanical properties and neovascularization over HAM in treatment of Achilles tendon injury in the early stage of healing.

A light-activated amnion wrap strengthens colonic anastomosis and reduces peri-anastomotic adhesions.

BACKGROUND AND OBJECTIVE: Colonic anastomotic failure is a dreaded complication, and multiple surgical techniques have failed to eliminate it. Photochemical tissue bonding (PTB) is a method of sealing tissue surfaces by light-activated crosslinking. We evaluated if a human amniotic membrane (HAM), sealed over the anastomotic line by PTB, increases the anastomotic strength. STUDY DESIGN: Sprague-Dawley rats underwent midline laparotomy followed by surgical transection of the left colon. Animals were randomized to colonic anastomosis by one of the following methods (20 per group): single-layer continuous circumferential suture repair (SR); SR with a HAM wrap attached by suture (SR+ HAM-S); SR with HAM bonded photochemically over the anastomotic site using 532 nm light (SR+ HAM-PTB); approximation of the bowel ends with only three sutures and sealing with HAM-PTB (3+ HAM-PTB). A control group underwent laparotomy alone with no colon resection (NR). Sub-groups (n = 10) were sacrificed at days 3 and 7 post-operatively and adhesions were evaluated. A 6 cm section of colon was then removed and strength of anastomosis evaluated by burst pressure (BP) measurement. RESULTS: A fourfold increase in BP was observed in the SR+ HAM-PTB group compared to suture repair alone (94 ± 3 vs. 25 ± 8 mm Hg, P < 0.0001) at day 3. At day 7 the burst pressures were 165 ± 40 and 145 ± 31 mm Hg (P = 1), respectively. A significant decrease in peri-anastomotic adhesions was observed in the SR+ HAM-PTB group compared to the SR group at both time points (P < 0.001). CONCLUSION: Sealing sutured colonic anastomotic lines with HAM-PTB increases the early strength of the repair and reduces peri-anastomotic adhesions. Lasers Surg. Med. 48:530-537, 2016. © 2016 Wiley Periodicals, Inc.

Improving Outcomes in Immediate and Delayed Nerve Grafting of Peripheral Nerve Gaps Using Light-Activated Sealing of Neurorrhaphy Sites with Human Amnion Wraps.

BACKGROUND: Photochemical tissue bonding uses visible light to create sutureless, watertight bonds between two apposed tissue surfaces stained with photoactive dye. When applied to nerve grafting, photochemical tissue bonding can result in superior outcomes compared with suture fixation. Our previous success has focused on immediate repair. It was the aim of this study to assess the efficacy of photochemical tissue bonding when performed following a clinically relevant delay. METHODS: Forty male Lewis rats had 15-mm left sciatic nerve gaps repaired with reversed isografts immediately (n = 20) or after a 30-day delay (n = 20). Repairs were secured using either suture or photochemical tissue bonding. Rats were killed after 150 days. Outcomes were assessed using monthly Sciatic Function Index evaluation, muscle mass retention, and nerve histomorphometry. Statistical analysis was performed using analysis of variance and the post hoc Bonferroni test. RESULTS: In both immediate and delayed groups, photochemical tissue bonding showed a trend toward greater recovery of Sciatic Function Index, but these results were not significant. The Sciatic Function Index was significantly greater when performed immediately. Significantly greater muscle mass retention occurred following photochemical tissue bonding in both immediate and delayed repairs. Values did not differ significantly between immediate and delayed groups. Histomorphometric recovery was greatest in the immediate photochemical tissue bonding group and poorest in the delayed suture group. Fiber diameter, axon diameter, myelin thickness, and G-ratio were not significantly different between immediate suture and delayed photochemical tissue bonding. CONCLUSIONS: Light-activated sealing of nerve grafts results in significantly better outcomes in comparison with conventional suture. The technique not only remains efficacious but may also help ameliorate the detrimental impacts of surgical delay.

Light-Activated Sealing of Nerve Graft Coaptation Sites Improves Outcome following Large Gap Peripheral Nerve Injury.

BACKGROUND: Nerve repair using photochemically bonded human amnion nerve wraps can result in superior outcomes in comparison with standard suture. When applied to nerve grafts, efficacy has been limited by proteolytic degradation of bonded amnion during extended periods of recovery. Chemical cross-linking of amnion before bonding may improve wrap durability and efficacy. METHODS: Three nerve wraps (amnion, cross-linked amnion, and cross-linked swine intestinal submucosa) and three fixation methods (suture, fibrin glue, and photochemical bonding) were investigated. One hundred ten Lewis rats had 15-mm left sciatic nerve gaps repaired with isografts. Nine groups (n = 10) had isografts secured by one of the aforementioned wrap/fixation combinations. Positive and negative control groups (n = 10) were repaired with graft and suture and no repair, respectively. Outcomes were assessed using sciatic function index, muscle mass retention, and histomorphometry. Statistical analysis was performed using analysis of variance and the post hoc Bonferroni test (p < 0.05). RESULTS: Cross-linking improved amnion durability. Photochemically bonded cross-linked amnion recovered the greatest sciatic function index, although this was not significant in comparison with graft and suture. Photochemically bonded cross-linked amnion recovered significantly greater muscle mass (67.3 ± 4.4 percent versus 60.0 ± 5.2 percent; p = 0.02), fiber diameter, axon diameter, and myelin thickness (6.87 ± 2.23 µm versus 5.47 ± 1.70 µm; 4.51 ± 1.83 µm versus 3.50 ± 1.44 µm; and 2.35 ± 0.64 µm versus 1.96 ± 0.47 µm, respectively) in comparison with graft and suture. CONCLUSION: Light-activated sealing of cross-linked human amnion results in superior outcomes when compared with conventional suture.

Light-activated sealing of skin wounds.

BACKGROUND AND OBJECTIVES: We have developed a light-activated technology for rapidly sealing skin surgical wounds called photochemical tissue bonding (PTB). The goals of this study were to evaluate parameters influencing PTB in order to optimize its clinical efficacy and to determine whether PTB can be used to seal wounds in moderately to highly pigmented skin. STUDY DESIGN/MATERIALS AND METHODS: Application of Rose Bengal (RB) followed by exposure to 532 nm was used to seal linear incisions (1.5 mm deep, 2 cm long) in lightly pigmented (Yorkshire) and darkly pigmented (Yucatan) swine skin. The force required to open the seal (the bonding strength) was measured by in situ tensiometry. Reflectance spectra, epidermal transmission spectra, and histology were used to characterize the skin. The relationships of RB concentration and fluence to bonding strength were established in Yorkshire skin. Surface temperature was measured during irradiations and cooling was used while sealing incisions in Yucatan skin. Monte Carlo simulations were carried out to estimate the effect of epidermal melanin on the power absorbed in the dermis at the incision interface. RESULTS: The lowest fluence, 25 J/cm(2), delivered at an irradiance of 0.5 W/cm(2) substantially increased the bonding strength (∼ 10-fold) compared to controls in Yorkshire swine skin. Increasing the fluence to 100 J/cm(2) enhanced bonding strength by a further 1.5-fold. Application of 0.1% RB for 2 minutes produced the greatest bonding strength using 100 J/cm(2) and limited the penetration of RB to an ∼ 50 µm band on the dermal incision wall. Reflectance spectra indicated that Yorkshire skin had minimal melanin and that Yucatan skin was a good model for highly pigmented human skin. In Yucatan skin, the bonding strength increased 1.7-fold using 0.1% RB and 200 J/cm(2) at 1.5 W/cm(2) with cooling and epinephrine. Monte Carlo simulation indicated that absorption of 532 nm light by epidermal melanin in dark skin decreased the power absorbed along the incision in the dermis by a factor of 2.7. CONCLUSIONS: These results suggest that in lightly pigmented skin the PTB treatment time can be shortened without compromising the bonding strength. Sealing incisions using PTB in moderately and highly pigmented skin will require a careful balance of irradiance and cooling.

Melanocytes are selectively vulnerable to UVA-mediated bystander oxidative signaling.

Long-wave UVA is the major component of terrestrial UV radiation and is also the predominant constituent of indoor sunlamps, both of which have been shown to increase cutaneous melanoma risk. Using a two-chamber model, we show that UVA-exposed target cells induce intercellular oxidative signaling to non-irradiated bystander cells. This UVA-mediated bystander stress is observed between all three cutaneous cell types (i.e., keratinocytes, melanocytes, and fibroblasts). Significantly, melanocytes appear to be more resistant to direct UVA effects compared with keratinocytes and fibroblasts, although melanocytes are also more susceptible to bystander oxidative signaling. The extensive intercellular flux of oxidative species has not been previously appreciated and could possibly contribute to the observed cancer risk associated with prolonged UVA exposure.

Prevention of capsular contracture with photochemical tissue passivation.

BACKGROUND: Capsular contracture is the most common complication following the insertion of breast implants. Within a decade, half of patients will develop capsular contracture, leading to significant morbidity and need for reoperation. There is no preventative treatment available and the recurrence rate remains high. Photochemical tissue passivation is a novel tissue-stabilization technique that results in collagen cross-linking. It can rapidly link collagen fibers in situ, preserving normal tissue architecture. By using this therapy to passivate the collagenous tissues of the implant pocket, the authors hope to prevent the development of pathogenic collagen bundles and subsequent capsule contracture. METHODS: Six-cubic centimeter tissue expanders were placed below the panniculus carnosus muscle along the dorsum of New Zealand white rabbits. Fibrin glue was instilled into each implant pocket to induce contracture. Treated pockets received photochemical tissue passivation by coating them with a photosensitizing dye and exposing the area to a 532-nm light. After 8 weeks, capsule tissue was harvested for histologic evaluation. RESULTS: Implant capsule thickness is the number one prognostic factor for contracture development. The authors demonstrated a 52 percent decrease in capsule thickness in the passivated group compared with controls. Photochemical tissue passivation resulted in fewer fibrohistiocytic cells and macrophages and in reduced synovial metaplasia and smooth muscle actin deposition. CONCLUSIONS: Photochemical tissue passivation significantly decreased both capsule thickness and smooth muscle actin deposition. It is a promising technique for preventing capsular contracture that can be performed at the time of initial surgery without a significant increase in procedure time.

Collagen cross-linking using rose bengal and green light to increase corneal stiffness.

PURPOSE: Photochemical cross-linking of corneal collagen is an evolving treatment for keratoconus and other ectatic disorders. We evaluated collagen cross-linking by rose bengal plus green light (RGX) in rabbit eyes and investigated factors important for clinical application. METHODS: Rose bengal (RB, 0.1%) was applied to deepithelialized corneas of enucleated rabbit eyes for 2 minutes. The diffusion distance of RB into the stroma was measured by fluorescence microscopy on frozen sections. RB-stained corneas were exposed to green (532-nm) light for 3.3 to 9.9 minutes (50-150 J/cm(2)). Changes in the absorption spectrum during the irradiation were recorded. Corneal stiffness was measured by uniaxial tensiometry. The spatial distribution of the stromal elastic modulus was assessed by Brillouin microscopy. Viable keratocytes were counted on H&E-stained sections 24 hours posttreatment. RESULTS: RB penetrated approximately 100 µm into the corneal stroma and absorbed >90% of the incident green light. RGX (150 J/cm(2)) increased stromal stiffness by 3.8-fold. The elastic modulus increased in the anterior approximately 120 µm of stroma. RB was partially photobleached during the 2-minute irradiation, but reapplication of RB blocked light transmission by >70%. Spectral measurements suggested that RGX initiated cross-linking by an oxygen-dependent mechanism. RGX did not decrease keratocyte viability. CONCLUSIONS: RGX significantly increases cornea stiffness in a rapid treatment (≅12 minutes total time), does not cause toxicity to keratocytes and may be used to stiffen corneas thinner than 400 µm. Thus, RGX may provide an attractive approach to inhibit progression of keratoconus and other ectatic disorders.