Luca Lancerotto

Necrotizing fasciitis: classification, diagnosis, and management.

Necrotizing fasciitis (NF), a life-threatening rare infection of the soft tissues, is a medical and surgical emergency. It is characterized by subtle, rapid onset of spreading inflammation and necrosis starting from the fascia, muscles, and subcutaneous fat, with subsequent necrosis of the overlying skin. Once suspected, immediate and extensive radical debridement of necrotic tissues is mandatory. Appropriate antibiotics and intensive general support avoid massive systemic diffusion of the infective process and are the key for successful treatment. However, early diagnosis is missed or delayed in 85% to 100% of cases in large published series: because of the lack of specific clinical features in the initial stage of the disease, it is often underestimated or confused with cellulitis or abscess. Mortality rates are still high and have shown no tendency to decrease in the last 100 years. Unfortunately, the prevalence of the disease is such that physicians rarely become sufficiently confident with NF to be able to proceed with rapid diagnosis and management. This review covers the literature published in MEDLINE in the period 1970 to December 31, 2010. Particular attention is given to the clinical and laboratory elements to be considered for diagnosis. A wide variety of diagnostic tools have been described to facilitate and hasten the diagnosis of NF, but the most important tool for early diagnosis still remains a high index of clinical suspicion.

Histological study of the deep fasciae of the limbs

The aim of this study is to analyse the deep fasciae of limbs in order to evaluate the collagen and elastic fibre arrangement and the types of innervation. Histological and immunohistochemical stains were performed in 72 specimens. The deep fascia of the limbs is a sheath presenting a mean thickness of 1mm, formed by two to three layers of parallel collagen fibre bundles. In the adjacent layers, they show different orientations. Each layer is separated from the adjacent one by loose connective tissue, permitting the sliding of the collagen layers. Nerve fibres were found in all specimens, while muscular fibres were evidenced only in one specimen. The described structure permits the fasciae of the limbs to have a strong resistance to traction, even when exercised in different directions. The capacity of the different collagen layers to glide one on the other could be altered in cases of overuse syndrome, trauma, or surgery.

Development of Mast Cells and Importance of Their Tryptase and Chymase Serine Proteases in Inflammation and Wound Healing

Mast cells (MCs) are active participants in blood coagulation and innate and acquired immunity. This review focuses on the development of mouse and human MCs, as well as the involvement of their granule serine proteases in inflammation and the connective tissue remodeling that occurs during the different phases of the healing process of wounded skin and other organs. The accumulated data suggest that MCs, their tryptases, and their chymases play important roles in tissue repair. While MCs initially promote healing, they can be detrimental if they are chronically stimulated or if too many MCs become activated at the same time. The possibility that MCs and their granule serine proteases contribute to the formation of keloid and hypertrophic scars makes them potential targets for therapeutic intervention in the repair of damaged skin.

Comparison of Transverse Carpal Ligament and Flexor Retinaculum Terminology for the Wrist

PURPOSE To investigate the macroscopic anatomy and histological characteristics of the transverse carpal ligament and the flexor retinaculum of the wrist and to investigate their anatomical relationships and define appropriate terminology. METHODS The volar regions of the wrists of 30 unembalmed subjects were examined by dissection and by histological and immunohistochemical staining. In vivo magnetic resonance imaging studies were also carried out on 10 subjects. RESULTS The dissection study showed that the antebrachial fascia at the volar aspect of the wrist presents a reinforcement. From a histological point of view, it is composed of 3 layers of undulated collagen fiber bundles. Adjacent layers show different orientations of the collagen fibers. Many nerve fibers and Pacini and Ruffini corpuscles were found in all specimens. Under this fibrous plane is another fibrous structure, placed transversely between the ulnar-sided hamate and pisiform bones, and the radial-sided scaphoid and trapezium bones. The deeper fibrous structure shows completely different histological characteristics, having parallel, thicker collagen fiber bundles and few nerve fibers. Magnetic resonance images confirm the presence of 2 clearly distinguished fibrous structures in the wrist, the first in continuity with the antebrachial fascia and the second located in a deeper plane between the hamate and scaphoid. CONCLUSIONS Two different fibrous structures with different histological characteristics are present in the volar wrist: the more superficial one is in continuity with the antebrachial fascia and could be considered its reinforcement; the deeper one is composed of strong lamina, with histological features similar to those of a ligament. For these reasons, we suggest that the term transverse carpal ligament should be used to indicate the fibrous lamina connecting the hamate and pisiform to the scaphoid and trapezium and that the term flexor retinaculum of the wrist should be abandoned because it does not correspond to any specific, autonomous structure.

External volume expansion increases subcutaneous thickness, cell proliferation, and vascular remodeling in a murine model.

Background: Fat grafting is a powerful tool for soft-tissue reconstruction; however, the science behind recipient bed preparation has not been thoroughly explored. External volume expansion using suction before fat grafting has been used clinically to improve reliability and consistency of graft survival. The authors developed a murine model to investigate the underlying mechanism of external volume expansion. Methods: The authors created an external volume expansion device using a soft-silicone dome connected to a vacuum source (25 mmHg) to treat the dorsum of mice, and the response was compared with treatment with an occlusive dressing. Treated areas were monitored with magnetic resonance imaging. Remodeling of microvasculature was studied with corrosion casting on day 7. Effects on tissue thickness, number of adipocytes, cell proliferation, and blood vessel density were analyzed at 28 days. Results: Macroscopic analysis showed tissue swelling at sites treated with the external volume expansion device by 21 days, without skin damage. On day 28, external volume expansion increased the thickness of the subcutaneous fat layer twofold, consistent with magnetic resonance imaging observations. The proliferation rate in the subcutaneous layer of expansion-treated areas increased twofold, with a net 2.2-fold increase in number of adipocytes in columns; remodeling of the vessels network occurred, with reorientation and increase of vessel diameters shown by corrosion casting and 1.9-fold augmentation of vessels density. Conclusions: External volume expansion applied to mouse integument induces highly proliferative and vascularized subcutaneous tissue. Recipient-site preparation using external volume expansion devices may be a promising tool to enhance cell and tissue engraftment.

Mechanisms of action of microdeformational wound therapy.

Microdeformational Wound Therapy (MDWT) is a class of medical devices that have revolutionized the treatment of complex wounds over the last 20 years. These devices, are a subset of Negative Pressure Wound Therapy (NPWT), in which there is a highly porous interface material placed between the wound and a semi-occlussive dressing and connected to suction. The porous interface material acts to deform the wound on a micro scale promoting cellular proliferation. These devices appear to significantly improve the speed of healing in many wounds, facilitate granulation tissue formation and reduce the complexity of subsequent reconstructive operations. The mechanisms through which such effects are obtained are beginning to be better understood through basic research and clinical trials. Further work in this field is likely to yield devices that are designed to treat specific wound types.

Neural potential of a stem cell population in the adipose and cutaneous tissues

Abstract Objective: A significant amount of recent interest has been focused on the possibility that adult human stem cells are a realistic therapeutic alternative to embryonic stem cells. Multipotent stem cells that have characteristics reminiscent of embryonic neural crest stem cells have been isolated from several postnatal tissues, including skin, gut, dental pulp and the heart, and are potentially useful for research and therapeutic purposes. However, their neurogenic potential, including their ability to produce electrophysiologically active neurons, is largely unexplored. In the present work, we investigated this issue with regard to skin-derived precursors (SKPs) and adipose-derived stem cells (ADSc) Methods: Adult stem cells isolated from skin and from adipose tissue derived from the same adult donor were treated with epidermal growth factor (EGF) and fibroblast growth factor 2 (FGF2). Neurospheres obtained were first expanded and evaluated in term of proliferative ability, and then their neuronal differentiation potential was analysed. Results: Adipose- and skin-derived neurospheres grew in suspension as spheres in the presence of the mitogens FGF2 and EGF. With this protocols, the spheres have been able to proliferate and to originate Schwann and glial-like cells. Discussion: In summary, we have demonstrated in this work that multipotent adult precursor cell can be isolated and expanded from two accessible adult tissue sources: skin and adipose tissue. The work described in this paper provides the framework for our attempts to use SKPs or ADSc as autologous adult stem cell population for cell replacement and discovery research.

Diffusion and Perfusion: The Keys to Fat Grafting

Background: Fat grafting is now widely used in plastic surgery. Long-term graft retention can be unpredictable. Fat grafts must obtain oxygen via diffusion until neovascularization occurs, so oxygen delivery may be the overarching variable in graft retention. Methods: We studied the peer-reviewed literature to determine which aspects of a fat graft and the microenvironment surrounding a fat graft affect oxygen delivery and created 3 models relating distinct variables to oxygen delivery and graft retention. Results: Our models confirm that thin microribbons of fat maximize oxygen transport when injected into a large, compliant, well-vascularized recipient site. The “Microribbon Model” predicts that, in a typical human, fat injections larger than 0.16 cm in radius will have a region of central necrosis. Our “Fluid Accommodation Model” predicts that once grafted tissues approach a critical interstitial fluid pressure of 9 mm Hg, any additional fluid will drastically increase interstitial fluid pressure and reduce capillary perfusion and oxygen delivery. Our “External Volume Expansion Effect Model” predicts the effect of vascular changes induced by preoperative external volume expansion that allow for greater volumes of fat to be successfully grafted. Conclusions: These models confirm that initial fat grafting survival is limited by oxygen diffusion. Preoperative expansion increases oxygen diffusion capacity allowing for additional graft retention. These models provide a scientific framework for testing the current fat grafting theories.

Quiescent Platelets Stimulate Angiogenesis and Diabetic Wound Repair

INTRODUCTION Platelets partake in hemostasis, wound healing, and tumor growth. Although platelet-rich-plasma (PRP) has been used in surgery for several years, its mechanism of action and application methods are still poorly characterized. MATERIALS AND METHODS A single unit of human platelets obtained by plateletpheresis was diluted in plasma and divided into three equal volumes. One volume was stored at room temperature as fresh platelets (RT), another volume was frozen by storage at -80 degrees C (FZ), and the third volume was frozen at -80 degrees C with 6% DMSO (FZ6). Plasma (PL) was used as control. Using flow cytometry, platelets were tested for platelet glycoprotein GPIb and annexin V binding, as survival and activation markers, respectively. Hemostatic function was assessed by thromboelastometry. In vivo, platelets were topically applied on 1 cm,(2) full-thickness wounds on db/db mice (n = 10/group) and healing was staged microscopically and macroscopically. RESULTS All platelet preparations showed hemostatic ability. RT platelets were GPIb positive (nonactivated-quiescent platelets) and stimulated angiogenesis by threefold, and cell proliferation by fourfold in vivo. FZ platelets were positive for annexin V, indicating activated platelets and, in vivo, increased only wound granulation. FZ6 platelets contained 30% nonactivated-quiescent and 50% activated platelets and stimulated granulation, angiogenesis, cell proliferation, and promoted re-epithelialization in vivo. CONCLUSIONS Platelets showed distinct mechanisms to induce hemostasis and wound healing. Quiescent platelets are required to induce angiogenesis in vivo. Platelets stored at room temperature and frozen with 6% DMSO and stored at -80 degrees C achieved optimal wound healing in diabetic mice.

Histological evolution of chronic wounds under negative pressure therapy

BACKGROUND Negative pressure wound therapy (NPT) has achieved widespread success in the treatment of difficult wounds. However, its effects are but partially explored, and investigations mostly concentrated at the wound-dressing interface; a detailed histological description of the evolution of wounds under NPT is still lacking. MATERIALS AND METHODS Subsequent punch biopsies of NPT-treated chronic wounds of human patients were analysed. Phenomena occurring in wounds were quantified by analysis of proliferating cells nuclear antigen (PCNA) (proliferating nuclei), CD31 (blood vessels), CD68p (macrophages) and CD45 (lymphocytes) stained slides. RESULTS Three layers were identified in day-0 wounds. Over time, under NPT, the layers behaved differently: the most superficial (1.5 mm) developed granulation tissue, constant in thickness, with high proliferation index, increased in blood vessels density and developed acute inflammation. Instead, the two deeper layers decreased in proliferation rate, maintained vessels density unchanged, were cleared of chronic inflammation and oedema and underwent progression towards stable tissue. DISCUSSION Indeed, while most research has focused on induction of superficial granulation tissue by NPT, deeper layers appear to be also affected, with relieving of chronic inflammation and tissue stabilisation. This may be an important and under-appreciated effect, playing a role in the known positive outcomes of NPT, such as better graft-taking rates.