Paula M. Farthing

Development of biodegradable electrospun scaffolds for dermal replacement

Our objective is to develop a synthetic biodegradable replacement dermal substitute for tissue engineering of skin and oral mucosa. Our in vivo criteria were that candidate scaffolds should allow surrounding cells to migrate fully into the scaffolds, enabling vasculogenesis and remodelling without invoking a chronic inflammatory response. We examined a total of six experimental electrospun polymer scaffolds: (1) poly-l-lactide (PLLA); (2) PLLA+10% oligolactide; (3) PLLA+rhodamine and (4-6) three poly(d,l)-lactide-co-glycolide (PLGA) random multiblock copolymers, with decreasing lactide/glycolide mole fractions (85:15, 75:25 and 50:50). These were evaluated for degradation in vitro up to 108 days and in vivo in adult male Wistar rats from 4 weeks to 12 months. In vivo, all scaffolds permitted good cellular penetration, with no adverse inflammatory response outside the scaffold margin and with no capsule formation around the periphery. The breakdown rate for each scaffold in vitro versus in vivo was similar, and an increase in the ratio of polyglycolide to polylactide correlated with an increase in breakdown rate, as expected. Scaffolds of PLLA were stable in vivo even after 12 months whereas scaffolds fabricated from PLGA 85:15 and 75:25 revealed a 50% loss of mass after 4 and 3 months, respectively. In vitro PLGA 85:15 and 75:25 scaffolds were able to support keratinocyte, fibroblast and endothelial cell growth and extracellular matrix production, with evidence of new collagen production after 7 days. In conclusion, the data supports the development of PLGA 85:15 and 75:25 electrospun polymer scaffolds as potential degradable biomaterials for dermal replacement.

Preliminary investigation of novel bone graft substitutes based on strontium–calcium–zinc–silicate glasses

Bone graft procedures typically require surgeons to harvest bone from a second site on a given patient (Autograft) before repairing a bone defect. However, this results in increased surgical time, excessive blood loss and a significant increase in pain. In this context a synthetic bone graft with excellent histocompatibility, built in antibacterial efficacy and the ability to regenerate healthy tissue in place of diseased tissue would be a significant step forward relative to current state of the art philosophies. We developed a range of calcium–strontium–zinc–silicate glass based bone grafts and characterised their structure and physical properties, then evaluated their in vitro cytotoxicity and in vivo biocompatibility using standardised models from the literature. A graft (designated BT109) of composition 0.28SrO/0.32ZnO/0.40 SiO2 (mol fraction) was the best performing formulation in vitro shown to induce extremely mild cytopathic effects (cell viability up to 95%) in comparison with the commercially available bone graft Novabone® (cell viability of up to 72%). Supplementary to this, the grafts were examined using the standard rat femur healing model on healthy Wister rats. All grafts were shown to be equally well tolerated in bone tissue and new bone was seen in close apposition to implanted particles with no evidence of an inflammatory response within bone. Complimentary to this BT109 was implanted into the femurs of ovariectomized rats to monitor the response of osteoporotic tissue to the bone grafts. The results from this experiment indicate that the novel grafts perform equally well in osteoporotic tissue as in healthy tissue, which is encouraging given that bone response to implants is usually diminished in ovariectomized rats. In conclusion these materials exhibit significant potential as synthetic bone grafts to warrant further investigation and optimisation.

ADRENOMEDULLIN HAS MITOGENIC EFFECTS ON HUMAN ORAL KERATINOCYTES : INVOLVEMENT OF CYCLIC AMP

The effects of the novel vasoactive regulatory peptide, adrenomedullin, on human oral keratinocytes was investigated. Adrenomedullin, acting via its specific receptor, stimulated a dose‐dependent increase in DNA synthesis, and, in addition, stimulated further changes in the cell cycle resulting in the proliferation of keratinocytes. When cells were incubated in the presence of increasing concentrations of adrenomedullin, there was a rapid and dose‐dependent rise in intracellular cyclic AMP levels. Stimulation of mitogenesis and cell proliferation in these cells were mimicked by the cell permeable cAMP analogue, di‐butyryl cAMP. Adrenomedullin‐stimulated mitogenesis was attenuated by the adenylyl cyclase inhibitor SQ22,536, but was unaffected by inhibitors of PKC, tyrosine kinase or the CGRP receptor antagonist, CGRP(8–37). These data identify adrenomedullin as a new mitogenic regulatory peptide of keratinocytes acting via the cAMP cascade.

Mechanisms of binding of cutaneous lymphocyte-associated antigen-positive and αeβ7-positive lymphocytes to oral and skin keratinocytes

Intraepithelial lymphocytes (IEL) utilize the integrin αeβ7 on their surface to bind to E‐cadherin on epithelial cells in the gut and breast. In oral mucosa and skin IEL express αeβ7 and the cutaneous lymphocyte‐associated antigen (CLA) but the mechanisms of adhesion of these subsets to keratinocytes are unknown. Levels of αeβ7 and CLA were up‐regulated on peripheral blood lymphocytes (PBL) by transforming growth factor‐β (TGF‐β) and interleukin‐12 (IL‐12), respectively, and both groups of lymphocytes adhered onto oral and skin keratinocytes. Adhesion of IL‐12‐activated PBL was totally abolished by anti‐lymphocyte‐associated function antigen type 1 (anti‐LFA‐1) antibodies but was unaffected by anti‐αeβ7 antibodies indicating that adhesion of the CLA‐positive subset is mediated via LFA‐1 interaction with intercellular adhesion molecule‐1 (ICAM‐1). Adhesion of TGF‐β‐activated PBL to E‐cadherin‐positive oral and skin keratinocytes was partially inhibited by anti‐αeβ7 antibodies but was unaffected by the blocking antibody E4.6 against E‐cadherin which detects the binding site for αeβ7‐positive lymphocytes in breast and gut epithelium. TGF‐β‐activated PBL also bound to an E‐cadherin‐negative oral keratinocyte cell line and adhesion was inhibited by anti‐αeβ7 antibodies. These results strongly suggest that in oral epithelium and epidermis αeβ7‐positive lymphocytes do not bind to E‐cadherin and there may be a novel second ligand for the αeβ7 integrin.

The pathology of oral cancer and precancer

Abstract Oral cancer is the sixth most common cancer worldwide and represents about 5.5% of all malignancies. In the Western world it is less common, but the incidence is increasing and the mortality rate has not improved for decades. Although most oral cancers probably arise in clinically normal mucosa some are preceded by a precancerous lesion which indicates an increased risk of cancer development at a particular site. The histopathologists role is to recognize pathological features which indicate high risk and to provide prognostic information from examination of excised tumours. The most common precancerous lesion is a white patch on the oral mucosa referred to as a leukoplakia. These show variable clinical features ranging from relatively innocuous flat white plaques to verruciform or red and white speckled lesions. A proportion of precancerous lesions show features of cytological atypia which are usually graded as mild, moderate or severe. The degree of epithelial dysplasia is a useful guide for the mangement of such lesions. 1 1Most oral malignancies are squamous cell carcinomas which are usually well differentiated and show similar features to squamous cell carcinomas elsewhere. Although a number of grading systems are available, of most practical value in routine diagnosis is an evaluation of the depth and pattern of invasion which has been shown to correlate most closely with tumour behaviour. Lesions with a diffuse or reticular pattern of invasion with small cords or single cells infiltrating the connective tissues have the worst prognosis. This infiltrative pattern also correlates with local lymph-node metastasis and when examining a neck dissection the histopathologist must pay particular attention to the level of nodal involvement in the neck and to extracapsular spread from involved nodes. Particular diagnostic difficulties may arise when variants of squamous cell carcinoma are encountered. These include anaplastic lesions, spindle cell carcinoma, adenosqumous carcinomas and basaloid squamous cell carcinoma. Occasionally benign lesions may show pseudoepitheliomatous hyperplasia which may be mistaken for malignancy.

Comparison of in vitro and in vivo bioactivity of SrO-CaO-ZnO-SiO2 glass grafts.

A range of calcium—strontium—zinc—silicate glass grafts are developed. Following characterization, their ability to form an apatite layer in simulated body fluid (SBF) is evaluated. Concurrently, their in vivo biocompatibility is determined. These glasses are incapable of forming an apatite layer in SBF. However, in vivo, each glass is well tolerated with new bone formation apparent in close apposition to implanted particles and no evidence of an inflammatory response. Such results are contrary to much of the literature and indicate that forecasting a materials ability to bond to bone based on SBF experiments may provide a false negative result.

Pyostomatitis vegetans and associated systemic disease : a review and two case reports

In this article two case reports of pyostomatitis vegetans are presented. Both cases were associated with inflammatory bowel disease and one case was also associated with liver dysfunction. It has recently been reported that there may be a link between liver dysfunction and pyostomatitis vegetans. The management of the condition is illustrated and the literature on the subject is reviewed.

Functional expression of the chemokine receptor XCR1 on oral epithelial cells.

Chemokines are chemoattractant cytokines which act on specific receptors and play an important role in leukocyte migration as well as physiological and pathological processes. We investigated the role of the chemokine receptor XCR1 and its ligand lymphotactin (Lptn/XCL1) in the regulation of oral epithelial cell behaviour. In vitro XCR1 mRNA and cell surface protein expression was detected in normal oral keratinocytes and oral squamous cell carcinoma cell lines. Lymphotactin mediated intracellular activation of the ERK1/2 signalling pathway and stimulated migration, invasion, and proliferation of all cells through XCR1. Oral cancer cells showed a greater response to lymphotactin than normal keratinocytes and a direct relationship between receptor expression and migration, invasion, and proliferation was observed. Exposure of normal keratinocytes to lymphotactin resulted in increased adhesion to fibronectin but not collagen and stimulated MMP‐2 and MMP‐9 but not MMP‐7 release, whereas exposure of cancer cells resulted in increased adhesion to both collagen and fibronectin and stimulated production of MMP‐2, MMP‐9, and MMP‐7. We observed XCR1 but not lymphotactin to be expressed by epithelial cells in normal oral mucosa in vivo, whilst both were expressed and up‐regulated in inflammatory oral disease and oral cancer including primary and metastatic disease. Lymphotactin mRNA and constitutive intracellular protein were detected in normal keratinocytes and oral cancer cell lines in vitro. These findings show that XCR1 and its ligand, lymphotactin, are expressed by oral epithelial cells and suggest that they play a role in regulating the behaviour of these cells. Copyright

Cytokine regulation of major histocompatibility complex antigen expression by human oral and skin keratinocytes

The expression, and cytokine modulation, of major histocompatibility complex (MHC) class I and class II molecules on oral and skin keratinocytes were compared in cell culture. Both cell types expressed class I, but not class II, constitutively. However, stimulation with interferon-gamma, but not interleukin-1 alpha, and -1 beta, tumour necrosis factor-alpha or lymphotoxin, induced increased expression of class I and de-novo expression of HLA-DR on both cell types. Oral keratinocytes differed from skin keratinocytes in that they exhibited greater sensitivity to interferon-gamma stimulation and higher stimulated expression of both class I and HLA-DR. In addition, interferon-gamma stimulated oral, but not skin, keratinocytes to express HLA-DP and -DQ. These observations suggest that, like skin keratinocytes, under certain conditions, oral keratinocytes may be able to act as antigen-presenting cells. This may be important in the initiation and progression of some immune-mediated mucocutaneous diseases. Moreover, differences in MHC expression may help to explain differences in the presentation of these diseases on the skin and oral mucosa.

Synovial sarcoma presenting as a parotid mass: case report and review of literature.

Synovial sarcoma is an unusual neoplasm of mesenchymal derivation, which is uncommon in the head and neck sites. In the parotid gland, it is most likely to be misdiagnosed as a myoepithelial, primary mesenchymal, or metastatic neoplasm.