OBJECTIVE Synovial fluid basic calcium phosphate (BCP) crystals are associated with severe joint degeneration accompanied by synovial hypertrophy. The metalloprotease 92 kDa gelatinase (MMP-9) has ...been implicated in the degradation of extracellular matrix in osteoarthritis, but the ability of BCP crystals to induce gelatinase in human fibroblasts or in adult porcine chondrocytes has not previously been studied. The hypothesis that the mitogenic response to BCP crystals is accompanied by induction and secretion of MMP-9 was studied. METHODS MMP-9 messenger RNA (mRNA) was detected by northern blot and reverse transcription-polymerase chain reaction (RT-PCR). Gelatinase secretion was identified by western blot and zymography of conditioned media. RESULTS BCP crystals caused a concentration dependent induction of MMP-9 mRNA accumulation and protein secretion in human fibroblasts but not in adult porcine chondrocytes. CONCLUSION BCP crystals induce MMP-9 production by HF but not adult porcine chondrocytes. Fibroblast MMP-9 may be an important mediator of the joint destruction associated with synovial fluid BCP crystals.
Cellular responses to whitlockite Ryan, L M; Cheung, H S; LeGeros, R Z ...
Calcified tissue international,
11/1999, Letnik:
65, Številka:
5
Journal Article
Recenzirano
Whitlockite crystals have been observed in both degenerating and normal articular cartilages. To determine their potential for inducing cartilage degeneration, we studied their ability to induce ...mitogenesis and synthesis and secretion of metalloproteases in vitro. Whitlockite crystals were found to stimulate cell proliferation and to stimulate synthesis and secretion of stromelysin and collagenase. However, they were less stimulatory than crystals that contained calcium (Ca) and phosphate without magnesium substitution for Ca. Whitlockite crystals elicit biologic cellular responses that suggest potential pathogenicity in arthritis, but are less potent than Ca phosphate crystals without magnesium.
We describe a 33-year-old woman with acute calcific periarthritis (ACP) of the interphalangeal joint of the thumb and review 42 reported cases of ACP involving the finger joints. A computer assisted ...literature search for reported cases of ACP involving the finger joints was performed. Clinical features of our case and those fulfilling the criteria for entry into this study were analyzed.
To investigate the potential therapeutic effects of inhibiting intracellular dissolution of basic calcium phosphate (BCP) crystals in tissue culture by raising lysosomal pH using bafilomycin A1, a ...specific vacuolar pump inhibitor.
45Ca-labeled crystals were used to demonstrate intracellular crystal dissolution in human foreskin fibroblasts (HF). Mitogenesis was evaluated using 3Hthymidine incorporation assays and cell counts. Northern blot and Western blot were used to study collagenase matrix metalloproteinase-1 (MMP1) mRNA accumulation and protein secretion, respectively.
Bafilomycin A1 inhibited intracellular dissolution of BCP crystals and caused a concentration-dependent inhibition of BCP crystal-induced mitogenesis. Doses of bafilomycin A1 which inhibited intracellular crystal dissolution and mitogenesis had no effect on BCP crystal-induced MMP1 mRNA accumulation or protein secretion.
Raising lysosomal pH to inhibit intracellular BCP crystal dissolution attenuates the proliferative response to BCP crystals in HF but does not prevent metalloprotease synthesis and secretion. The therapeutic potential of lysosomotropic agents for preventing joint destruction in BCP crystal deposition disease is limited.
We describe, for the first time, hydrogel-forming microneedle arrays prepared from "super swelling" polymeric compositions. We produced a microneedle formulation with enhanced swelling capabilities ...from aqueous blends containing 20% w/w Gantrez S-97, 7.5% w/w PEG 10,000 and 3% w/w Na2CO3 and utilised a drug reservoir of a lyophilised wafer-like design. These microneedle-lyophilised wafer compositions were robust and effectively penetrated skin, swelling extensively, but being removed intact. In in vitro delivery experiments across excised neonatal porcine skin, approximately 44 mg of the model high dose small molecule drug ibuprofen sodium was delivered in 24 h, equating to 37% of the loading in the lyophilised reservoir. The super swelling microneedles delivered approximately 1.24 mg of the model protein ovalbumin over 24 h, equivalent to a delivery efficiency of approximately 49%. The integrated microneedle-lyophilised wafer delivery system produced a progressive increase in plasma concentrations of ibuprofen sodium in rats over 6 h, with a maximal concentration of approximately 179 µg/ml achieved in this time. The plasma concentration had fallen to 71±6.7 µg/ml by 24 h. Ovalbumin levels peaked in rat plasma after only 1 hour at 42.36±17.01 ng/ml. Ovalbumin plasma levels then remained almost constant up to 6 h, dropping somewhat at 24 h, when 23.61±4.84 ng/ml was detected. This work represents a significant advancement on conventional microneedle systems, which are presently only suitable for bolus delivery of very potent drugs and vaccines. Once fully developed, such technology may greatly expand the range of drugs that can be delivered transdermally, to the benefit of patients and industry. Accordingly, we are currently progressing towards clinical evaluations with a range of candidate molecules.
Phosphoinositides
are lipid second messengers known to be important for many cellular processes in yeast, including actin cytoskeletal organization, vesicle transport, and cell wall assembly. In ...plant cells, studies on phosphoinositide phosphatases and kinases suggest that phosphoinositides are involved in the regulation of actin cytoskeletal organization, cell wall synthesis, and cell morphogenesis. It is hypothesized that phosphoinositides may regulate the transport of vesicles carrying cell wall biosynthetic enzymes and wall components, thereby influencing cell wall synthesis and cell morphogenesis.
The design of a non-viral gene delivery vehicle capable of delivering and releasing a functional nucleic acid cargo intracellularly remains a formidable challenge. For systemic gene therapy to be ...successful a delivery vehicle is required that protects the nucleic acid cargo from enzymatic degradation, extravasates from the vasculature, traverses the cell membrane, disrupts the endosomal vesicles and unloads the cargo at its destination site, namely the nucleus for the purposes of gene delivery. This manuscript reports the extensive investigation of a novel amphipathic peptide composed of repeating RALA units capable of overcoming the biological barriers to gene delivery both in vitro and in vivo. Our data demonstrates the spontaneous self-assembly of cationic DNA-loaded nanoparticles when the peptide is complexed with pDNA. Nanoparticles were <100nm, were stable in the presence of serum and were fusogenic in nature, with increased peptide α-helicity at a lower pH. Nanoparticles proved to be non-cytotoxic, readily traversed the plasma membrane of both cancer and fibroblast cell lines and elicited reporter-gene expression following intravenous delivery in vivo. The results of this study indicate that RALA presents an exciting delivery platform for the systemic delivery of nucleic acid therapeutics.
Articular cartilage contains any ectoenzyme activity, NTP-PPH, which is capable of generating PPi from NTP substrates. The PPi generated is from the cleavage of the alpha-beta pyrophosphate bond of ...NTP and does not result from the effects of NTP catabolites. NTP-PPH activity is expressed on human skin fibroblasts in culture and is significantly increased in subjects with CPPD deposition. In addition, cultured fibroblasts from subjects with CPPD disease have higher intracellular PPi concentrations compared to cells from normals and patients with OA. These results support the hypothesis that alterations in PPi metabolism provide the metabolic basis for CPPD deposition.
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