Ilyas Khan | Cardiff University (original) (raw)

Papers by Ilyas Khan

Trends in genetics: TIG, Jan 1, 1994

PCR amplification followed by single-strand conformation polymorphism (SSCp) analysis is a well-e... more PCR amplification followed by single-strand conformation polymorphism (SSCp) analysis is a well-established method for screening for mutations in particular segments of DNA I. PCR amplification may be hampered by the generation of nonspecific products, particulady in sequences that are GC-rich. Specificity can be improved by including either DMSO, formaz:~ide 2 or the nucleotide analogue 7-deaza-2'..dGTP (Ref. 3) in the reaction. We have found that the presence of 3:1 7-deaza-2'-dGTP:dGTP greatly improves the specificity of PCR amplification of certain segments of the promoter regions of the human IGF2 gene. However, analysis of the PCR products by SSCP resulted in the formation of fuzzy, indistinct bands on gel electrophoresis (see .

Methods in Molecular …, Jan 1, 2007

Necrosis and apoptosis have been demonstrated in articular cartilage in response to trauma and di... more Necrosis and apoptosis have been demonstrated in articular cartilage in response to trauma and disease. However, cell death in articular cartilage may also be thought of as a scale of cell death culminating in secondary necrosis with the failure to remove apoptotic cells from the tissue. The in situ detection of cell death is an important technique in studying articular cartilage as it most closely resembles the in vivo situation. The methods described here involve the use of light microscopy and electron microscopy in conjunction with fluorescent and biochemical methods to correctly ascertain the type of cell death that has occurred.

… Engineering Part A, Jan 1, 2009

Experimental wounding of articular cartilage results in cell death at the lesion edge. The object... more Experimental wounding of articular cartilage results in cell death at the lesion edge. The objective of this study was to investigate whether inhibition of this cell death results in enhanced integrative cartilage repair. Bovine articular cartilage discs (6 mm) were incubated in media containing inhibitors of necrosis (Necrostatin-1, Nec-1) or apoptosis (Z-VAD-FMK, ZVF) before cutting a 3 mm inner core. This core was left in situ to create disc/ring composites, cultured for up to 6 weeks with the inhibitors, and analyzed for cell death, sulfated glycosaminoglycan release, and tissue integration. Creating the disc/ring composites resulted in a significant increase in necrosis. ZVF significantly reduced necrosis and apoptosis at the wound edge. Nec-1 reduced necrosis. Both inhibitors reduced the level of wound-induced sulfated glycosaminoglycan loss. Toluidine blue staining and electron microscopy of cartilage revealed significant integration of the wound edges in disc/ring composites treated with ZVF. Nec-1 improved integration, but to a lesser extent. Push-out testing revealed that ZVF increased adhesive strength compared to control composites. This study shows that treatment of articular cartilage with cell death inhibitors during wound repair increases the number of viable cells at the wound edge, prevents matrix loss, and results in a significant improvement in cartilage-cartilage integration.

Genomics, Jan 1, 2000

The SON gene, which maps to human chromosome 21q22.1-q22.2, encodes a novel regulatory protein.

Osteoarthritis and Cartilage, Jan 1, 2009

Objective: Articular cartilage contains mesenchymally derived chondroprogenitor cells that have t... more Objective: Articular cartilage contains mesenchymally derived chondroprogenitor cells that have the potential to be used for stem cell therapy. The aim of this study was to characterise the growth kinetics and properties of in vitro expanded cloned chondroprogenitors and determine if critical determinants of the progenitor phenotype were maintained or lost in culture.

Arthritis & …, Jan 1, 2001

PloS one, Jan 1, 2010

Background: Articular cartilage displays a poor repair capacity. The aim of cell-based therapies ... more Background: Articular cartilage displays a poor repair capacity. The aim of cell-based therapies for cartilage defects is to repair damaged joint surfaces with a functional replacement tissue. Currently, chondrocytes removed from a healthy region of the cartilage are used but they are unable to retain their phenotype in expanded culture. The resulting repair tissue is fibrocartilaginous rather than hyaline, potentially compromising long-term repair. Mesenchymal stem cells, particularly bone marrow stromal cells (BMSC), are of interest for cartilage repair due to their inherent replicative potential. However, chondrocyte differentiated BMSCs display an endochondral phenotype, that is, can terminally differentiate and form a calcified matrix, leading to failure in long-term defect repair. Here, we investigate the isolation and characterisation of a human cartilage progenitor population that is resident within permanent adult articular cartilage.

Molecular …, Jan 1, 2006

Cultured HOSE cells exhibited extensive fluorescent dyecoupling and connexin43 (Cx43) expression;... more Cultured HOSE cells exhibited extensive fluorescent dyecoupling and connexin43 (Cx43) expression; other connexins were not detected. The ovarian adenocarcinoma cell lines had little dye-coupling or connexin expression. Deletions and rearrangements of the Cx43 gene were not detected by Southern blotting in the carcinoma lines. N 6 ,2Ј-O-dibutyryladenosine 3Ј,5Ј-cyclic monophosphate and alltrans-retinoic acid inhibited cell proliferation, but did not enhance GJIC or Cx43 expression. Surface epithelial cells of benign ovaries expressed Cx43, but this expression was barely detectable in ovarian serous cystadenocarcinomas. Thus, normal HOSE cells had extensive GJIC and Cx43 expression whereas ovarian carcinoma cells had less and cAMP and retinoic acid did not change these, although both agents inhibited cell growth.

Journal of …, Jan 1, 2006

Arguably, the gold standard of biological repair of articular cartilage lesions is autologous cho... more Arguably, the gold standard of biological repair of articular cartilage lesions is autologous chondrocyte transplantation. Although the clinical outcomes appear to range between good and excellent in most cases, there are, nevertheless, both clinical and biological challenges that remain to improve rehabilitation and clinical outcome. One of the major biological problems relates to tissue integration of the reparative tissue into the host tissue at a predictable level. Often within a single lesion, varying degrees of integration can be observed from total integration through to non-integration as one passes through the defect. Here we briefly review some of the literature relating to this problem and include some of our own data drawn from questions we have posed about the biological nature of cartilage/cartilage integration. The nature and status of the tissue that comprises the wound lesion edge is central to tissue integration, and controlling aspects of trauma and free-radical-induced cell death together with matrix synthesis are identified as two components that require further investigation. Interestingly, there appears to be a limited ability of chondrocytes to be able to infiltrate existing cartilage matrices and even to occupy empty chondrocyte lacunae. Proliferation as a result of blunt and sharp trauma shows differential responses. As expected, blunt trauma induces a greater proliferative burst than sharp trauma and is more widespread from the lesion edge. However, in the case of sharp trauma, the basal cells enter proliferation before surface zone chondrocytes, which is not the case in blunt wounds. Regulation of these and associated processes will be necessary in order to devise strategies that can predict successful integration in biological repair procedures.

…, Jan 1, 2009

Objective. To analyse the heterogeneity at the single-cell level of human mesenchymal progenitor ... more Objective. To analyse the heterogeneity at the single-cell level of human mesenchymal progenitor cells from SM. Methods. Cell populations were enzymatically released from the knee joint synovium of adult human individuals. Single cell-derived clonal populations were obtained by limiting dilution and serially passaged to determine growth rates. Phenotypic analysis was carried out by flow cytometry. Replicative senescence was assessed by the senescence-associated -galactosidase staining. Telomere lengths were determined semiquantitatively by Southern blotting. Telomerase activity was measured using a real-time quantitative telomerase repeat amplification procedure. Culture-expanded clonal populations were subjected to in vitro differentiation assays to investigate their mesenchymal multipotency.

Clinical …, Jan 1, 2001

Cartilage has a poor reparative capacity although it is unclear as to what extent this may be dep... more Cartilage has a poor reparative capacity although it is unclear as to what extent this may be dependent on age or maturation. In the current study, the cellular responses of chondrocytes to experimental wounding in vitro using embryonic, immature, and mature cartilage have been compared. In all cases, the response was consistent (a combination of cell death that included apoptosis and proliferation). The speed of response varied in terms of cell death with embryonic cartilage showing the most rapid response and mature cartilage showing the slowest response. Intrinsic repair as assessed by the ability to heal the lesion was not detected in any of the culture systems used. It was concluded that the poor repair potential of cartilage is not maturation dependent in the systems studied.

Arthritis & …, Jan 1, 2008

Methods. MSC populations were derived from adult synovium and periosteum. Phenotype analysis was ... more Methods. MSC populations were derived from adult synovium and periosteum. Phenotype analysis was performed by fluorescence-activated cell sorting and real-time reverse transcriptase-polymerase chain reaction (RT-PCR). Telomere lengths were determined by Southern blot analysis. In vitro osteogenesis was assessed quantitatively by measurements of alkaline phosphatase activity and calcium deposits. To investigate bone formation in vivo, MSCs were seeded onto osteoinductive scaffolds and implanted subcutaneously in nude mice. Bone was assessed by histology, and the human origin investigated by in situ hybridization for human Alu genomic repeats. Quantitation was achieved by histomorphometry and real-time RT-PCR for human osteocalcin. Analysis at the single-cell level was performed with clonal populations obtained by limiting dilution. Multiple regressions were used to explore the incremental predictive value of the markers.

Nucleic acids research, Jan 1, 1993

Arthritis & …, Jan 1, 2006

Methods. Cell populations were enzymatically released from the periosteum of the proximal tibia o... more Methods. Cell populations were enzymatically released from the periosteum of the proximal tibia obtained from adult human donors and then expanded in monolayer. Single-cell-derived clonal populations were obtained by limiting dilution. Culture-expanded periosteal cell populations were tested for their growth potential and for expression of conventional markers of MSCs and were subjected to in vitro assays to investigate their multilineage potential. To assess their multipotency in vivo, periosteal cells were injected into a regenerating mouse tibialis anterior muscle for skeletal myogenesis or were either seeded into an osteoinductive matrix and implanted subcutaneously into nude mice for osteogenesis or implanted in a joint surface defect under a periosteal flap into goats for chondrogenesis. Cell phenotypes were analyzed by histochemistry and immunohistochemistry and by reverse transcriptionpolymerase chain reaction for the expression of lineagerelated marker genes.

Journal of cell …, Jan 1, 2004