Topical vanadate enhances the repair of median laparotomy incisions.

BACKGROUND: There are over two million laparotomies performed in the United States each year with an incisional hernia rate between 2% and 11%. A total of 100,000 ventral hernia repairs are undertaken each year with recurrences as high as 50%. MATERIALS AND METHODS: Full thickness midline fascia incisions from the xiphoid to the pubic symphysis were made in rats. The fascia and/or muscular layer was sutured closed and a gel with 300 µM of sodium orthovanadate or saline was placed over the suture line with the skin closed over it. On day 10, 1-cm strips from the superior, middle, and inferior regions of the abdominal wall were tested for breaking strength and processed for histology. RESULTS: The mean wound breaking strength of vanadate-treated wounds was 18.6 ± 2.7 N compared with 9.4 ± 3.6 N for controls (P < 0.0001). Similar quantities of granulation tissue were deposited in treated and control wounds. Fine green birefringence patterns, characteristic of immature connective tissue, were seen in control samples viewed with polarized light. In contrast, vanadate-treated wounds showed thick yellow-orange birefringence patterns characteristics of more mature connective tissue. Using α-smooth muscle actin immunostaining, myofibroblasts were prominent in control incisions, but few were identified in vanadate-treated incisions. CONCLUSIONS: In rat laparotomy wounds, a single application of vanadate increases wound breaking strength, through enhanced connective tissue organization. These combined data suggest topical application of vanadate immediately after fascial closure will increase wound strength, possibly reducing hernia recurrences in the repaired abdominal wall.

Modulatory effects of connexin-43 expression on gap junction intercellular communications with mast cells and fibroblasts.

The influence of mast cells upon aberrant wound repair and excessive fibrosis has supportive evidence, but the mechanism for these mast cell activities is unclear. It is proposed that heterocellular gap junction intercellular communication (GJIC) between fibroblasts and mast cells directs some fibroblast activities. An in vitro model was used employing a rodent derived peritoneal mast cell line (RMC-1) and human dermal derived fibroblasts. The influence of the expression of the gap junction channel structural protein, connexin 43 (Cx-43) on heterocellular GJIC, the expression of microtubule ß-tubulin and microfilament α smooth muscle actin (SMA) were investigated. The knockdown of Cx-43 by siRNA in RMC-1 cells completely blocked GJIC between RMC-1 cells. SiRNA knockdown of Cx-43 within fibroblasts only dampened GJIC between fibroblasts. It appears Cx-43 is the only expressed connexin (Cx) in RMC-1 cells. Fibroblasts express other Cxs that participate in GJIC between fibroblasts in the absence of Cx-43 expression. Heterocellular GJIC between RMC-1 cells and fibroblasts transformed fibroblasts into myofibroblasts, expressing α SMA within cytoplasmic stress fibers. The knockdown of Cx-43 in RMC-1 cells increased ß-tubulin expression, but its knockdown in fibroblasts reduced ß-tubulin expression. Knocking down the expression of Cx-43 in fibroblasts limited αSMA expression. Cx-43 participation is critical for heterocellular GJIC between mast cells and fibroblasts, which may herald a novel direction for controlling fibrosis.

Fibroblast expression of α-smooth muscle actin, α2ß1 integrin and αvß3 integrin: influence of surface rigidity.

Open wound contraction necessitates cell and connective tissue interactions, that produce tension. Investigating fibroblast responses to tension utilizes collagen coated polyacrylamide gels with differences in stiffness. Human foreskin fibroblasts were plated on native type I collagen-coated polyacrylamide gel cover slips with different rigidities, which were controlled by bis-acrylamide concentrations. Changes in alpha smooth muscle actin (αSMA), α2ß1 integrin (CD49B) and αvß3 integrin (CD-51) were documented by immuno-histology and Western blot analysis. Cells plated on rigid gels were longer, and expressed αvß3 integrin and αSMA within cytoplasmic stress fibers. In contrast, cells on flexible gels were shorter, expressed α2ß1 integrin and had fine cytoskeletal microfilaments without αSMA. Increased tension changed the actin makeup of the cytoskeleton and the integrin expressed on the cell's surface. These in vitro findings are in agreement with the tension buildup as an open wound closes by wound contraction. It supports the notion that cells under minimal tension in early granulation tissue express α2ß1 integrin, required for organizing fine collagen fibrils into thick collagen fibers. Thicker fibers create a rigid matrix, generating more tension. With increased tension cytoskeletal stress fibers develop that contain αSMA and αvß3 integrin that replaces α2ß1 integrin, consistent with cell switching from collagen to non-collagen proteins interactions.

Demonstrating collagen tendon fibril segments involvement in intrinsic tendon repair.

Severed tendons can undergo regenerative healing, intrinsic tendon repair. Fibrillogenesis of chick tendon involves "collagen fibril segments" (CFS), which are the building blocks of collagen fibers that make up tendon fascicles. The CFS are 10.5 micron in length, composed of tropocollagen monomers arranged in parallel arrays. Rather than incorporating single tropocollagen molecules into growing collagen fibers, incorporating large CFS units is the mechanism for generating collagen fibers. Is intrinsic tendon repair through the reestablishment of tendon embryogenesis? Gentamicin treated 10-day-old chick embryo tendons released CFS were fluorescently tagged with Rhodamine (Rh). Organ cultured severed 14-day-old embryo tendon explants received Rh tagged CFS. At day 4 auto fluorescent tagged CFS were identified at the severed tendon ends by fluorescent microscopy. Accumulation of fluorescent tagged CFS was exclusively localized to the severed ends of tendon explants. Parallels between collagen fiber growth during embryonic fibrillogenesis and tendon repair reveal CFS incorporation is responsible for collagen fibers growth. CFS incorporation into frayed collagen fibers from severed tendons is the proposed mechanism for intrinsic tendon repair, which is an example of regenerative repair.

Differences in the mechanism of collagen lattice contraction by myofibroblasts and smooth muscle cells.

Both rat derived vascular smooth muscle cells (SMC) and human myofibroblasts contain α smooth muscle actin (SMA), but they utilize different mechanisms to contract populated collagen lattices (PCLs). The difference is in how the cells generate the force that contracts the lattices. Human dermal fibroblasts transform into myofibroblasts, expressing α-SMA within stress fibers, when cultured in lattices that remain attached to the surface of a tissue culture dish. When attached lattices are populated with rat derived vascular SMC, the cells retain their vascular SMC phenotype. Comparing the contraction of attached PCLs when they are released from the culture dish on day 4 shows that lattices populated with rat vascular SMC contract less than those populated with human myofibroblast. PCL contraction was evaluated in the presence of vanadate and genistein, which modify protein tyrosine phosphorylation, and ML-7 and Y-27632, which modify myosin ATPase activity. Genistein and ML-7 had no affect upon either myofibroblast or vascular SMC-PCL contraction, demonstrating that neither protein tyrosine kinase nor myosin light chain kinase was involved. Vanadate inhibited myofibroblast-PCL contraction, consistent with a role for protein tyrosine phosphatase activity with myofibroblast-generated forces. Y-27632 inhibited both SMC and myofibroblast PCL contraction, consistent with a central role of myosin light chain phosphatase.

When the Smad signaling pathway is impaired, fibroblasts advance open wound contraction.

In wound healing transforming growth factor ß1 (TGFß1), utilizing the Smad signaling pathway, advances connective tissue deposition, the transformation of fibroblasts into myofibroblasts and wound contraction. The compound SB-505124 disrupts the Smad signaling pathway by blocking activin receptor-like kinase phosphorylation of select Smad signaling proteins. Four full thickness excisional square 2×2 cm wounds were made on the rat dorsum. On day 2, the pair of wounds on the left received 1 µM SB-505124 in gel, and the pair on the right, controls, received gel alone. Wounds were covered with nonocclusive dressings and treated redressed daily for 4 days. No differences in day 14 wound sizes between treatment groups were found. H&E stained sections revealed increased cell density in SB-505124 treated wounds. Polarized light microscopy showed collagen fiber bundles birefringence intensity and organization were equivalent between treatment groups. Myofibroblast populations, identified by α-smooth muscle actin staining, were the norm in controls but absent in SB-505124 treated wounds, which was confirmed by Western blot analysis. Blocking the Smad signaling pathway diminished connective tissue deposition and generated a deficiency in myofibroblast numbers, but wound contraction was unimpaired. The absence of myofibroblasts may be related to the blocking of the Smad signaling pathway or it may be related to the generation of less tension in treated wounds, related to reduce deposited connective tissue. These findings support the notion that wound contraction does not require the generation of myofibroblast contractile forces, but rather the organization of newly deposited collagen fiber bundles by forces related to fibroblast locomotion.

COL1A1 oligodeoxynucleotides decoy: biochemical and morphologic effects in an acute wound repair model.

Type I collagen is an integral component of granulation tissue and scar, that is highly dependent on TGFß1, a member of a pro-fibrotic family of cytokines, for its promotion and deposition. Blocking COL1A1 gene transcription obstructs type I collagen synthesis, hindering the progress of granulation tissue deposition and fibrosis. Local injections of a double stranded oligodeoxynucleotide (dsODN) decoy, containing the TGFß1 regulatory element that is located in the distal promoter of the COL1A1 gene, were investigated in a rat polyvinyl alcohol (PVA) sponge granulation tissue implant model. The effects on the granulation tissue deposition by dsODN decoy therapy were evaluated by the synthesis of types I and III collagens as well as ED-A (cellular) fibronectin. Fluorescently labeled dsODN was used to identify the distribution of the decoy molecules in the sponge implant relative to the observed histological effects. Morphological alterations in cells and changes in the organization of connective tissue were documented and evaluated. Collagen levels were reduced by half in implants treated with 10 nM dsODN decoy compared to scrambled dsODN-treated implants. Histologically, dsODN decoy treated implants had an increased cellular density without a corresponding increase in deposited connective tissue. Polarized light birefringence pattern of Sirius red-stained sections showed less collagen fibers accumulating between fibroblasts. The highest concentration of fluorescently labeled dsODN was identified within the interior margin of sponge implants, correlating to increased cellular density and an altered birefringence patterns. In conclusion, 10 nM dsODN decoy therapy reduced collagen deposition and altered the organization of granulation tissue, supporting its potential as a localized anti-fibrotic therapy for limiting fibrotic conditions.

The role of vascular endothelial growth factor in wound healing.

BACKGROUND: A chronic wound is tissue with an impaired ability to heal. This is often a consequence of one of the following etiologies: diabetes, venous reflux, arterial insufficiency sickle cell disease, steroids, and/or pressure. Healing requires granulation tissue depending on epithelialization and angiogenesis. Currently no growth factor is available to treat patients with impaired healing that stimulates both epithelialization and angiogenesis. The objective is to review is the multiple mechanisms of vascular endothelial growth factor (VEGF) in wound healing. MATERIALS AND METHODS: The authors reviewed the literature on the structure and function of VEGF, including its use for therapeutic angiogenesis. Particular attention is given to the specific role of VEGF in the angiogenesis cascade, its relationship to other growth factors and cells in a healing wound. RESULTS: VEGF is released by a variety of cells and stimulates multiple components of the angiogenic cascade. It is up-regulated during the early days of healing, when capillary growth is maximal. Studies have shown the efficacy of VEGF in peripheral and cardiac ischemic vascular disease with minimal adverse effects. Experimental data supports the hypothesis that VEGF stimulates epithelialization and collagen deposition in a wound. CONCLUSION: VEGF stimulates wound healing through angiogenesis, but likely promotes collagen deposition and epithelialization as well. Further study of the molecule by utilizing the protein itself, or novel forms of delivery such as gene therapy, will increase its therapeutic possibilities to accelerate closure of a chronic wound.

Influence of vanadate on migrating fibroblast orientation within a fibrin matrix.

Treating rats with vanadate, a nonspecific inhibitor of protein tyrosine phosphatases, optimizes the uniform packing of collagen fiber bundles in wound granulation tissue and doubles wound breaking strength in rat incisional wounds. The speculation is vanadate optimizes the packing of collagen fiber bundles through the orientation of newly arrived wound fibroblasts in the fibrin clot filling the defect. Segments of 14 day chick embryo tendons were placed on fibrin clots and maintained in organ culture with and without 30 microM vanadate. On day 7 explants were examined histologically and biochemically. Tendon fibroblast outgrowth from untreated explants migrated in a random fashion, while fibroblasts from vanadate-treated explants migrated out in linear arrays. Fibroblasts were elongated by 20% form vanadate treated explant compared to controls. Myosin ATPase, required for optimal cell motility, is optimized by the phosphorylation of its myosin light chain (MLC). Western blot analysis of lysates from the fibroblasts that migrated into the fibrin showed vanadate increased MLC-P levles. These findings support the notion that vanadate promotes the deposition of regular, parallel collagen fiber bundles by advancing the orientation of fibroblasts in parallel linear arrays early in the wound repair process.

Tissue stretch decreases soluble TGF-beta1 and type-1 procollagen in mouse subcutaneous connective tissue: evidence from ex vivo and in vivo models.

Transforming growth factor beta 1 (TGF-beta1) plays a key role in connective tissue remodeling, scarring, and fibrosis. The effects of mechanical forces on TGF-beta1 and collagen deposition are not well understood. We tested the hypothesis that brief (10 min) static tissue stretch attenuates TGF-beta1-mediated new collagen deposition in response to injury. We used two different models: (1) an ex vivo model in which excised mouse subcutaneous tissue (N = 44 animals) was kept in organ culture for 4 days and either stretched (20% strain for 10 min 1 day after excision) or not stretched; culture media was assayed by ELISA for TGF-beta1; (2) an in vivo model in which mice (N = 22 animals) underwent unilateral subcutaneous microsurgical injury on the back, then were randomized to stretch (20-30% strain for 10 min twice a day for 7 days) or no stretch; subcutaneous tissues of the back were immunohistochemically stained for Type-1 procollagen. In the ex vivo model, TGF-beta1 protein was lower in stretched versus non-stretched tissue (repeated measures ANOVA, P < 0.01). In the in vivo model, microinjury resulted in a significant increase in Type-1 procollagen in the absence of stretch (P < 0.001), but not in the presence of stretch (P = 0.21). Thus, brief tissue stretch attenuated the increase in both soluble TGF-beta1 (ex vivo) and Type-1 procollagen (in vivo) following tissue injury. These results have potential relevance to the mechanisms of treatments applying brief mechanical stretch to tissues (e.g., physical therapy, respiratory therapy, mechanical ventilation, massage, yoga, acupuncture).

Oligodeoxynucleotide decoy therapy blocks type 1 procollagen transcription and the prolyl hydroxylase beta subunit translation.

Persistent transforming growth factor-beta1 (TGF-beta1) exposure to lungs increases type 1 collagen synthesis and deposition resulting in excess fibrosis which leads to morbidity and possibly death. We now report using human embryonic lung fibroblasts in the presence of TGF-beta1, a novel double-stranded (ds) DNA decoy with phosphorothioate (PT) linkages, containing the TGF-beta cis-element found in the distal promoter region of the COL1A1 gene which silences COL1A1 gene expression. In a cell-free protein translation system, we have previously reported that collagen synthesis was inhibited by disulfide isomerase, the prolyl-4-hydroxylase (P-4-H) beta subunit. By comparative proteomics dsdecoy therapy increased the levels of disulfide isomerase, the P-4-H beta subunit. These findings taken together support the notion that the dsdecoy inhibits type 1 collagen synthesis at both the transcriptional and translational levels.

A review of fibroblast-populated collagen lattices.

Bell's introduction of the fibroblast-populated collagen lattice (FPCL) has facilitated the study of collagen-cell interactions. As a result of the numerous modifications of the casting of FPCLs, the in vivo applications of these in vitro findings have been confusing. Here experimental FPCL contraction findings are viewed in regard to three proposed mechanisms responsible for lattice contraction. The cellular mechanisms responsible for generating FPCL contraction are cell contraction, cell tractional forces related to cell locomotion, and initial cell elongation and spreading.

The synergism of age and db/db genotype impairs wound healing.

Both diabetes and advanced age have been implicated in delaying wound repair. However, the contribution of age alone has not been shown clinically to significantly impair the ability to heal. To determine the contribution of age and db/db genotype multiple wound healing parameters were determined in young db/db mice, aged db/db mice, age-matched non-db/db control and wild-type C57BL/6 mice. Biomechanical properties (breaking load and tensile stiffness), epithelialization, and collagen deposition were determined for the four groups of mice 14 days after wounding with suture-closed incisional wounds. While neither hyperglycemia nor age alone caused impairment in biomechanical properties, the combination of age and db/db genotype resulted in a 36% reduction in stiffness and a 42% reduction in breaking load, when compared to young control mice, suggesting poor quality of healing. Statistically significant differences in the volume of granulation tissue deposited within the wound site were also observed, with the aged db/db mice displaying more than any other group, suggesting greater dermal loss from the dermal edges of incisional wounds in aged db/db mice, suggesting that the combination of age and diabetes act synergistically to impair healing in mice with type 2 diabetes. Interestingly, the impairment occurs independently of the prevailing glycemia, supporting the hypothesis that diabetes in synergy with advanced age has downstream effects, leading to further impairment, necessitating initiation of early and aggressive intervention in elderly patients with diabetic foot ulcers.

Understanding experimental biology of skin equivalent: from laboratory to clinical use in patients with burns and chronic wounds.

A major breakthrough in burn wound care was the early excision of the burn and its immediate coverage with a skin autograft. A search for a skin-graft substitute began to reduce the autografting-related trauma at the donor site. One entry was skin equivalence, which contains 3 components: (1) living fibroblasts, suspended in (2) a native collagen matrix, the surface of which is covered with (3) viable keratinocytes. The tissue-cultured dermal fibroblasts are derived from human foreskin. The fibroblasts are grown in cell culture dishes as a monolayer and are retrieved by limited trypsin digestion. The fibroblast suspension is mixed with serum-supplemented culture medium and native acid-soluble collagen. The entire mixture, called a dermal equivalent, is placed in a bacteriological Petri dish before transfer to a 37 degrees C incubator. The collagen rapidly polymerizes, trapping cells in the dermal equivalent. During the initial 4 hours, fibroblasts elongate and spread, causing a decrease in the thickness of the dermal equivalent. After 6 hours, the dermal equivalent undergoes a decrease in diameter as a consequence of the reorganization of the collagen. A freshly isolated suspension of human skin-derived keratinocytes is seeded on the surface of a several-day-old floating dermal equivalent. The keratinocytes proliferate, covering the surface of the dermal equivalent. The keratinocytes deposit basement membranes beneath them and undergo epidermal cell differentiation, leading to the formation of a basal layer beneath differentiated cell layers. Both cell populations retain viability and release cell factors that have a positive effect on wound closure. The placement of skin equivalence within a chronic wound may share structural attributes with a skin graft, but its function is to accelerate closure.

Is the tendon embryogenesis process resurrected during tendon healing?

The process of embryonic tendon development, including the nature and purpose of collagen fibril segments, is reviewed. It is proposed that tendon fibrillogenesis of repair is related to the fibrillogenesis of tendon embryonic development. The assembly of collagen fibril segment units into longer fibers occurs on the surface of tendon fibroblasts in embryonic tendon development. The biochemist's view of tendon healing, whereby the spontaneous polymerization of tropocollagen monomers regenerates lost tendon collagen fibers, needs to be reconsidered. Furthermore, the importance of direct fibroblast involvement in collagen fiber reassembly during tendon healing needs to be studied in tendon intrinsic regenerative repair.

Dupuytren's disease: physiologic changes in nodule and cord fibroblasts through aging in vitro.

The pathogenesis of the fibrotic disease Dupuytren's contracture remains unclear. The disease process includes two structurally distinct fibrotic elements, the nodule and the cord. It has been proposed that as the disease progresses, nodules develop into cords. To corroborate that hypothesis, the authors took advantage of cultured fibroblast differences found between gap junction intercellular communication and fibroblast-populated collagen lattice contraction. Paired fibroblast cell lines of nodules and cords derived from four patients with Dupuytren's disease were maintained in culture for at least eight passages. The presence of gap junction intercellular communication in nodule- and cord-derived fibroblasts was documented and reported as a coupling index. The contraction of free-floating nodule- or cord-derived collagen lattices was also documented and reported. Early passage (passage 4) cord-derived fibroblasts showed a significant increase in coupling index compared with passage 4 nodule-derived fibroblasts (4.0 +/- 0.4 versus 2.5 +/- 0.3, respectively), where p < or = 0.01. However, late passage (passage 8) nodule- and cord-derived fibroblasts were equivalent in their coupling index (4.1 +/- 0.4 versus 4.4 +/- 0.4, respectively). Early passage nodule-derived fibroblast-populated collagen lattices contracted by 64 percent, whereas late passage nodule-derived lattices showed less contraction, at only 40 percent. Early and late passage cord-derived lattices contracted 46 and 37 percent, respectively. All nodule- and cord-derived cell lines were statistically equivalent at lattice contraction by passage 8. These in vitro studies support the hypothesis that fibroblasts derived from Dupuytren's contracture nodules change their phenotype after undergoing repeated cell passage, acquiring a cord-like fibroblast phenotype. Dupuytren's nodules represent the early, active form of fibrosis in which cells are more proliferative, better at fibroblast-populated collagen lattice contraction, and display less gap junction intercellular communication. The speculation is that alterations in gap junction intercellular communication may be involved in the progression of Dupuytren's nodules to cords as the disease progresses.

Pulp nonfiction: microscopic anatomy of the digital pulp space.

The volar pad of the fingertip provides a very stable yet sensitive surface that gives the hand the ability to pinch and grasp. The focus of this study was to advance understanding of the anatomical features of the digital pulp space. The unusual features of the fingertip pulp space include prominent collagen fiber cords and a branching continuous fine vasculature. Prominent collagen fiber cords radiating out from beneath the epidermal basement membrane are like the cords of a parachute, which directly attach to the periosteum of the distal phalanx. Those collagen fiber cords are responsible for the firm attachment of the fingertip to the distal phalanx. There is a fine patent vasculature within the pulp space. Also contained in the capsule are numerous lobules of fat, which contribute to some elasticity of the fingertip. Principles of treatment for injuries or infections of the digital pulp should attempt to preserve this anatomical construct so that the firmness and vascular supply of the fingertip are maintained and not disrupted.

Clinical efficacy and mechanism of bilayered living human skin equivalent (HSE) in treatment of diabetic foot ulcers.

UNLABELLED: Bilayered living human skin equivalent (HSE) consists of cultured keratinocytes residing on the surface of a fibroblast-populated collagen lattice. Although HSE is FDA-approved for treatment of diabetic foot and venous stasis ulcers, its clinical efficacy remains limited, because the molecular mechanisms underlying its therapeutic effect are not fully understood. It is, therefore, often applied mistakenly as a skin graft. In this report, we delineate a mechanism of HSE biological effect and consequent optimal clinical use in accelerating closure of diabetic foot ulcers. EXPERIMENTAL: HSE was grafted onto nude mice and the release of various growth factors was evaluated by reverse transcription-polymerase chain reaction (RT-PCR) and immunochemistry. Clinical: HSE was grafted onto 11 consecutive patients with diabetes who had 13 non-ischemic foot ulcers and healing was measured as time to 100% closure (e.g., no drainage and 100% epithelialized). EXPERIMENTAL: HSE cellular components were determined to express 15 different growth factors/cytokine genes known to promote wound healing. Histological evidence from the nude mice showed that the collagen component of HSE underwent remodeling within the first seven days of grafting. Clinical: All diabetic foot ulcers healed in 31.8 12.4 days. Local release of a unique combination of 15 growth factors expressed by HSE keratinocyte and fibroblast components generates closure of diabetic foot ulcers. HSE should be applied with the same surgical conditions for a skin graft (i.e., no cellulitis, no drainage, and negligible bacteria). We hypothesize that bilayered HSE generates its effect by way of the local synthesis and release of multiple growth factors in specific combination and concentration, which improves the impaired reparative process of chronic wounds.

Healing of elderly patients with diabetic foot ulcers, venous stasis ulcers, and pressure ulcers.

Although elderly patients have physiologic impairments in wound healing, their wounds should be expected to heal with the same frequency of closure as those in younger populations, albeit at a slower rate. However, compared to the general population, the elderly population has a higher incidence of chronic wounds: diabetic foot ulcers, pressure ulcers, and venous stasis ulcers. Experimental and clinical data indicate physiologically impaired healing is characterized by decreased angiogenesis and synthesis of critical growth factors. Further, compared to younger populations, the elderly have a higher rate of mortality associated with specific morbidities, such as sepsis and acute respiratory distress. As these morbidities may develop directly from the wound, early intervention is mandated. In this report, 40 consecutive elderly patients (65-102 years old) with chronic wounds were analyzed. All patients were provided the same treatment protocol and healing was defined as 100% epithelization and no drainage. Despite the wounds presenting in a nonhealing and/or infected state, 73% of these chronic wounds in elderly patients healed. This suggests that elderly patients with diabetic foot ulcers, pressure ulcers, and venous stasis ulcers close their wounds at a similar frequency as younger patients. Therefore, early intervention and comprehensive treatment that includes safe topical therapies, in addition to growth factors and cellular therapy used for chronic wounds, ensure these patients will be spared the morbidities of pain, amputation, osteomyelitis, and even death. We hypothesize that if all elderly patients with chronic wounds are provided early treatment, morbidities (e.g., amputation, sepsis, pain) and associated costs will decrease.

Mast cells prevent dexamethasone-induced cell death of cultured fibroblasts: relationship to gap junctional intercellular communications.

BACKGROUND: Dexamethasone, a common therapy for reducing hypertrophic scar, sometimes fails. However, in cell culture, all dexamethasone-treated fibroblasts die. In co-cultures, gap junction intercellular communications between mast cells and fibroblasts promote profibrotic activities. Does the co-culture of mast cells with fibroblasts prevent dexamethasone-induced fibroblast death? METHODS: Survival of fibroblasts co-cultured with RMC-1 cells, a rat mast cell line, receiving dexamethasone was studied. RMC-1 cells pretreated with a secretagogue that degranulated mast cells and/or with a long-acting gap junction intercellular communications inhibitor were compared to untreated RMC-1 cells co-cultured with fibroblasts and dexamethasone. RESULTS: Fibroblasts alone treated with dexamethasone all died in 3 hours. Fibroblasts co-cultured with intact RMC-1 cells or with degranulated RMC-1 cells in dexamethasone all survived. No fibroblasts survived, co-cultured with RMC-1 cells unable to form gap junction intercellular communications with fibroblasts. CONCLUSIONS: Dexamethasone-treated fibroblasts, forming gap junction intercellular communications with mast cells, may explain why dexamethasone therapy sometimes fails. Gap junction intercellular communications between scar mast cells and fibroblasts or myofibroblasts apparently blocks the death of these cell populations. Preventing gap junction intercellular communications between mast cells and fibroblasts by including anti-gap junction intercellular communication agents may enhance the effectiveness of steroid therapy in treating excessive scarring.