Quantitative analysis of gene expression in human articular chondrocytes in monolayer culture

Human articular chondrocytes (HACs) were isolated from cartilage samples of normal joints and cultivated in monolayer for 56 days. Collagen type I, II, IX, aggrecan, and versican expression was quantified by real-time polymerase chain reaction (PCR). The expression of the genes was highly time-dependent and changed over the culture time. Collagen type I was not present at the beginning of the culture and increased 100-fold during the culture time. Collagen type II and IX expression was found during the entire culture period and decreased more than 100-fold. Aggrecan expression was downregulated 100-fold whereas versican expression increased 10 times. Indices of cell differentiation, defined as ratios of collagen type II to I (CII/I), collagen type IX to I (CIX/I), and aggrecan to versican (Agg/Ver), were significantly higher at the beginning of the culture and decreased over the culture period. The CII/I and CIX/I ratios formed three phases, with a plateau at the beginning, followed by a transition phase and a steady state at the end of the culture time, whereas the Agg/Ver ratio declined constantly. The accurate quantitative assessment of extracellular matrix gene expression in samples of chondrocytes taken from monolayer culture can be used to monitor chondrocyte metabolism as well as changes in the cell differentiation status.     

Effect of vinpocetine on retrograde axoplasmic transport

Vinpocetine, a derivate of vincamine, is widely used in the clinical pharmacotherapy of cerebral circulatory diseases. Herewith we report on a novel effect of vinpocetine: inhibition of retrograde axoplasmic transport of nerve growth factor (NGF) in the peripheral nerve. Blockade of retrograde transport of NGF results in transganglionic degenerative atrophy (TDA) in the segmentally related ipsilateral superficial spinal dorsal horn, which is characterized by depletion of the marker enzymes fluoride-resistant acid phosphatase (FRAP) and thiamine monophosphatase (TMP). At the same time, pain-related neuropeptides such as substance P (SP) and calcitonin gene-related peptide (CGRP), are depleted from lamina I-III from the segmentally related, ipsitateral Rolando substance of the spinal cord. On the basis of these experiments it is suggested that vinpocetine may result in a locally restricted decrease of nociception, that might be useful in clinical treatment of intractable pain. Pilot self-experiments support this assumption.     

A pilot study on the antibodies to HHV-6 variants and HHV-7 in CSF of MS patients

In the possible role for human herpesviruses (HHV) in the pathogenesis of multiple sclerosis (MS) neither clear distinction between the two variants of HHV-6, nor the involvement of HHV-7 have been described. Therefore, we quantitated HHV-6 variant specific and HHV-7 reacting antibodies in the CSF of 13 patients with MS or other neurological disorders by ELISA. Predominance in the positivity of IgG (67%) and IgM (44%) to HHV-6B over that of IgG (44%) with no detectable IgM to HHV-6A, and no antibodies to HHV-7 were found in the CSF of MS patients. None of these antibodies were found in the CSF of controls. This suggests that, intrathecal chronic active or primary HHV-6B infection might contribute to MS progression, while the local effects of HHV-6A and HHV-7 seem to be less important.     

Severe hypoaldosteronism due to corticosterone methyl oxidase type II deficiency in two boys: metabolic and gas chromatography-mass spectrometry studies

Infection-triggered, life-threatening salt-loss and hyperkalaemia developed in two male infants with wasting, inappropriately low plasma aldosterone concentrations and elevated plasma renin activity. The presumptive diagnosis of a defective terminal step in aldosterone biosynthesis was made by the presence of large amounts of 11-dehydrotetrahydrocorticosterone and its 18-hydroxylated metabolite (18-OH-THA), free 18-hydroxycorticosterone (18-OH-B) and 18-hydroxytetra-hydrocorticosterone in the urine of both patients. The diagnosis of corticosterone methyl oxidase type II (CMO II) deficiency was confirmed by an elevated urinary 18-OH-THA to tetrahydroaldosterone ratio in one boy and by an elevated plasma 18-OH-B to aldosterone ratio in the other boy. Unknown steroids responsible for the salt-loss were not identified. Sodium supplementation but not short-term high dose oral 9-fluorcortisol (FF) normalized the hyponatraemia in one patient, in whom sodium (Na⁺)/potassium (K⁺) co-transport was decreased. Both patients eventually received long-term FF treatment to prevent impairment of longitudinal growth caused by chronic salt-loss. The diagnosis of CMO II deficiency should always be confirmed by elevated precursor-product ratios in urine or plasma, using radioimmunoassays with prior chromatographic separation. Metabolic studies as the short-term response of serum Na⁺ to high dose FF may not be helpful in differentiating aldosterone biosynthetic defects from endorgan resistance to mineralocorticoids.Dedicated to Professor Dr. Walter Teller, on the occasion of his 60th birthdayPresented in part at the 27th Annual Meeting of the European Society for Paediatric Endocrinology, Copenhagen, Denmark, June 1988     

Do meta-analyses reveal time-dependent differences between the clinical outcomes achieved by microfracture and autologous chondrocyte implantation in the treatment of cartilage defects of the knee?

Purpose

To test the hypothesis that autologous chondrocyte implantation (ACI) has a better treatment effect than microfracture (MF), and increasing superiority over the years, when performed under similar patient-specific and defect-specific conditions.

Methods

We scanned four electronic databases for controlled clinical trials or controlled prospective observational studies. We conducted random-effects meta-analyses of equivalent data using standardized mean differences as the outcome measure of choice at 1, 2, and 5-year follow-up. We assessed heterogeneity with the I ² index and publication bias with funnel plots and Kendall’s tests.

Results

Our literature search revealed six study populations (nine papers) which satisfied our eligibility criteria. Overall, 399 patients aged between 16 and 60 years with 1–10 cm² chondral defects were available. The MF and the ACI study groups were well matched regarding patient baseline characteristics. For all papers, microfracture was performed according to Steadman, whereas three generations of ACI were applied. When all were combined, non-significant superiority of ACI over MF was revealed; surprisingly, this superiority decreased over the years. However, our meta-analyses combining solely second and third-generation ACI revealed significant standardized differences, becoming smaller over the years, but always representing a large effect. Nevertheless, our approximate estimate of the difference between the treatment effects provoked by second and third-generation ACI and by MF is not indicative of clinically relevant superiority of ACI over MF at 5-year follow-up.

Conclusions

Both series of meta-analyses (combining either all ACI modifications or solely the second and third generations of ACI) suggest that the treatment effects resulting from ACI and MF converge over the years.     

Thromboembolic complications after spinal surgery in trauma patients

Background Deep venous thrombosis (DVT) and pulmonary embolism (PE) may be significant complications following spinal surgery. The incidence rate ranges from 0.5% to 2.5% in patients with symptomatic thromboembolic disease and up to 15% in patients with non-symptomatic thrombotic complications. We determined the incidence of symptomatic thromboembolism after spinal surgery in patients with postoperative systemic prophylaxis and investigated general and specific risk factors for development of this disease.

Patients and methods We analyzed the clinical records of 978 patients who had undergone surgery of the spine because of trauma and who had been admitted to our level-I trauma center between 1980 and 2004. Spinal procedures included anterior and/or posterior spinal fusion, video-assisted thoracoscopic fusion, and spinal decompression. Symptomatic thromboembolic disease was diagnosed when patients showed significant clinical signs or symptoms of DVT or PE. In cases of DVT, diagnosis was confirmed by duplex scan of the lower limbs; in cases of PE, diagnosis was confirmed by CT-scanning of the thorax or at post mortem.

Results The incidence rate of symptomatic thromboembolic complications was 2.2% (n 22). 17 patients showed clinical signs of deep venous thrombosis, with 4 of them developing pulmonary embolism subsequently. The other 5 patients developed pulmonary embolism without prior clinical signs of deep venous thrombosis. 6 patients died because of thromboembolic disease. Thromboembolic complications were more frequent in older patients and among males, as well as in patients with regular tobacco consumption and obesity. Thromboembolic complications were also seen more frequently in patients with surgical procedures at the lumbar spine, in patients with anterior spinal fusion, and in those with motor deficits in the lower extremities.

Interpretation We found a rather low rate of clinically significant thromboembolic complications after spinal surgery because of trauma, compared to the results reported in the literature. Level of spinal surgery, surgical approach, and motor deficits in the lower extremities were identified as specific risk factors for DVT or PE. Age, sex, obesity and regular smoking were identified as general risk factors.