1,25(OH)2D3 and dihydrotestosterone interact to regulate proliferation and differentiation of epiphyseal chondrocytes

Growth plate chondrocytes are affected by 1,25(OH)₂D₃ and androgens, which may critically interact to regulate proliferation and differentiation during the male pubertal growth spurt. We investigated possible interactions of 1,25(OH)₂D₃ and the non-aromatizable androgen dihydrotestosterone (DHT) in primary chondrocyte cultures from young male rats. DHT and 1,25(OH)₂D₃ independently stimulated DNA synthesis and cell proliferation in a dose-dependent manner with maximally effective doses of [10^-8 M] and [10^-12 M], respectively. Both DHT and 1,25(OH)₂D₃ stimulated the expression and release of IGF-I, and the proliferative effects of each hormone were prevented by an IGF-I antibody. DHT and 1,25(OH)₂D₃ increased messenger RNAs (mRNAs) of their cognate receptors and of IGF-I receptor mRNA (IGF-I-R). 1,25(OH)₂D₃ also stimulated mRNA of the androgen receptor (AR), whereas DHT did not affect mRNA of the vitamin-D receptor (VDR). Coincubation with both steroid hormones did not stimulate receptor mRNAs more than either hormone alone. The proliferative effects of DHT and 1,25(OH)₂D₃ were completely inhibited by simultaneous incubation with both hormones, despite potentiation of IGF-I synthesis. In contrast, both hormones synergistically stimulated cell differentiation as judged by alkaline phosphatase activity, collagen X mRNA, and matrix calcification in long-term experiments. We conclude that DHT and 1,25(OH)₂D₃ interact with respect to chondrocyte proliferation and cell differentiation. The proliferative effects of both hormones are mediated by local IGF-I synthesis. Simultaneous coincubation with both hormones blunts the proliferative effect exerted by either hormone alone, in favor of a more marked stimulation of cell differentiation.     

Tamoxifen-inducible Cre-recombination in articular chondrocytes of adult Col2a1-CreER(T2) transgenic mice

OBJECTIVE: To determine the specificity and efficiency of the tamoxifen (TM)-induced Cre-recombination in articular chondrocytes of adult Col2a1-CreER(T2) transgenic mice. METHODS: Col2a1-CreER(T2) transgenic mice were bred with Rosa26 reporter mice. Two-week-old Col2a1-CreER(T2);R26R mice were administered TM for 5 days and were sacrificed 1 and 6 months after TM induction. X-Gal staining was performed. RESULTS: Efficient Cre-recombination is achieved in adult articular chondrocytes 1 and 6 months after TM induction. CONCLUSION: Our findings demonstrate that the Col2a1-CreER(T2) transgenic mouse model is a valuable tool to target genes specifically expressed in articular chondrocytes in a temporally controlled manner in adult mice.     

15d-PGJ(2) is acting as a 'dual agent' on the regulation of COX-2 expression in human osteoarthritic chondrocytes

The PGD(2) metabolite 15-deoxy-delta12,14 PGJ(2) (15d-PGJ(2)), a potent peroxisome proliferator-activated receptor gamma (PPARgamma) activator, has recently received attention for its potential antiinflammatory effects, but its effect on the cyclooxygenase-2 (COX-2) production is still under debate. We investigated the effect of 15d-PGJ(2) on COX-2 and prostaglandin E(2) (PGE(2)) production in the absence or the presence of interleukin-1beta (IL-1beta) in human osteoarthritic chondrocytes.Data showed that, as expected, IL-1beta induced both COX-2 and PGE(2) production. The addition of 15d-PGJ(2) completely blocked (by 93%) the IL-1beta-induced PGE(2) synthesis, whereas COX-2 level was only partially reduced (by 72%). Interestingly in the absence of any COX-2 inducer, 15d-PGJ(2) up-regulated COX-2 expression without concomitant elevation of PGE(2) synthesis. This study showed that the PPARgamma agonist, 15d-PGJ(2), exerts a dual effect on COX-2 production. The mechanisms by which 15d-PGJ(2) favors COX-2 production will be discussed.     

羟基磷灰石-软骨细胞复合物的构建

目的探讨以块状多孔羟基磷灰石（ＨＡ）作为软骨细胞培养的载体,构建一种促进骨愈合的植入材料的可行性。方法采取成年雄性ＳＤ大鼠肋软骨细胞,以３ｍｍ×３ｍｍ×４ｍｍ大小的块状多孔ＨＡ为载体培养软骨细胞;培养３天和７天后通过扫描电镜观察ＨＡ表面和中矢横断面孔隙中有无细胞生长,同时将培养２天的细胞爬片以α１（Ⅱ）ｃＤＮＡ片段为探针进行原位杂交,求证培养的细胞是否软骨细胞。结果细胞爬片原位杂交染色强阳性,证实是软骨细胞,且ＨＡ表面和中矢横断面孔隙内壁上均有细胞生长,随培养时间增加细胞数明显增加。结论ＨＡ可以作为细胞培养的贴壁底物,软骨细胞可以长入多孔ＨＡ内部的孔隙中。     

Considerations about angiolymphoid hyperplasia with eosinophilia (ALHE) with regard to a case localized in the penis

We report the case of a 27 year old man who presented a nodule in the ventral face of the penis, which increased in volume and turned painful during erection so a vascular lesion was suspected, later confirmed with a Doppler study. A complete excision of the lesion was carried out with local anaesthesia. The pathological specimen was informed as "angiolimphoid hyperplasia with eosinophilia" in its intravascular form. This is the third communication of this kind of lesion in the penis but no other adopted the intravascular variant like the present case. We discuss about this exceptional entity and make review of the most relevant literature.     

Plasmapheresis with a substitution solution of human serum protein (5%) versus plasmapheresis with a substitution solution of human albumin (5%) in patients suffering from autoimmune diseases

Therapeutic plasma exchange (TPE) has been used extensively for over 2 decades to treat a variety of autoimmune and congenital diseases and is now widely accepted. The primary objective of this study was to compare the clinical efficacy of two plasma exchange preparations, human serum protein (HSP) and human albumin (HA). Twenty-four patients in the following disease categories underwent TPE using either HSP (Biseko, 5%) or HA (5%): systemic lupus erythematosus, 8; glomerulonephritis, 8; myasthenia gravis, 2; Guillain-Barré-syndrome, 2; recurrent iritis, 1; pemphigoid, 1; uveitis, 1; and vascular retinitis, 1. There was no statistically significant difference in the average number of TPEs needed in the HSP group (13.5) and HA (13.8) measured over the first 6 weeks of treatment. The secondary parameters, in particular the immunological parameters IgG and IgA, provided evidence that plasma exchange with HSP may have some advantages over HA, and confirmatory studies in a larger group of patients are indicated. Adverse events during TPE occurred in both the HAS group (4 patients) and the HA group (4 patients). However, patients in the HSP group were older (12.3 years), were suffering from more complicated autoimmune diseases, and the number of occasions (days) on which these were reported (6 days) was less than in the HA group (11 days). One patient in the HA group died from septic-toxic circulatory collapse on Day 49 due to an infection with resistant strains of Staphylococcus aureus. Infections in other patients did not occur; all showed considerable improvement in their symptoms and completed the study in good general condition.     

Value of scintigraphic examination methods with 99mTechnetium in injuries of the epiphyseal cartilage (author's transl)

The value of scintigraphic examinations of injuries involving the epiphyseal plate is to be seen to ensure the correct diagnosis of the lesions type I and type V according to Salter and Harris. Further on the results of these scintigraphic examinations allow a better planning of the therapeutic procedure. Experimental findings in rabbits succeeded to predict the seriousness and an early prognosis. The disturbances of the epiphyseal blood flow can be diagnosed by 99m Tc-O4 labeled erythrocytes and 99m Tc-O4 labeled albumin microspheres. 99m Tc-MDP skeletal scintigraphy gives the information at which time the repair is done and full weight bearing is possible.     

藻酸钙复合自体软骨细胞修复羊膝关节负重区软骨缺损的实验研究

[目的]以中国山羊为动物模型,观察藻酸钙复合自体软骨细胞修复膝关节负重区软骨缺损的可行性。[方法]取羊肩关节软骨,分离、培养软骨细胞,蕃红"O"、 Giemsa及Ⅱ型胶原免疫组织化学染色对其进行鉴定。将自体软骨细胞与藻酸钙凝胶复合,修复山羊股骨髁负重区全层软骨缺损(直径6 mm),实验分为四组:(1)缺损旷置组:缺损内未植入任何组织;(2)骨膜覆盖组:自体骨膜覆盖缺损区;(3)藻酸钙+骨膜组:凝胶植入软骨缺损区,并用自体骨膜覆盖;(4)藻酸钙+细胞+骨膜组:藻酸钙复合自体软骨细胞植入软骨缺损区,自体骨膜覆盖;分别于手术后3、6个月取材,通过大体观察及组织学评分检测修复效果。[结果]软骨细胞复合物蕃红"O"、 Giemsa染色及Ⅱ型胶原免疫组化染色结果均为阳性,将藻酸钙凝胶-软骨细胞复合物用于羊负重区关节面软骨缺损修复,从大体观察和组织学评分进行比较,发现各组均有不同程度的组织修复,藻酸钙+细胞+骨膜组效果最好,与其他组差异有统计学意义(P<0.05)。[结论]藻酸钙凝胶-软骨细胞复合物结合自体骨膜覆盖,可较好修复山羊膝关节负重区软骨缺损。     

In Vivo Treatment with Calcitriol (1,25(OH)2D3) Reverses Age-Dependent Alterations of Intestinal Calcium Uptake in Rat Enterocytes

The vitamin D endocrine system has been involved in the impairment of intestinal calcium absorption during aging. Alterations in the nongenomic mechanism of calcitriol (1,25-dihydroxy-vitamin D₃; [1,25(OH)₂D₃] have been recently evidenced. In enterocytes isolated from aged rats, 1,25(OH)₂D₃ stimulation of Ca²⁺ channels through the cAMP/PKA pathway is blunted. We have now investigated whether in vivo administration of calcitriol to senescent rats reverses the absence of hormonal effects in isolated intestinal cells. In enterocytes from 20–24-month-old rats given 1,25(OH)₂D₃ for 3 days (30 ng/100 g bw/day), calcitriol (10⁻¹⁰ M, 3–5 minutes) stimulated Ca²⁺ uptake and intracellular cAMP to the same degree and protein quinase A (PKA) activity to a lesser degree than in enterocytes from young animals. Significantly higher basal levels of cAMP and PKA detected in enterocytes from old rats were not affected by prior injection of animals with 1,25(OH)₂D₃. When the aged rats were injected with 25(OH)D₃, similar Ca²⁺ influx, cAMP, and PKA responses to in vitro stimulation with calcitriol were obtained. 1,25(OH)₂D₃-dependent changes in Ca²⁺ uptake by enterocytes from both young and old rats treated with calcitriol were totally suppressed by the cAMP antagonist Rp-cAMPS, whereas the response to the agonist Sp-cAMPS was markedly depressed in aged animals. These results suggest that intestinal resistance to nongenomic 1,25(OH)₂D₃ stimulation of duodenal cell Ca²⁺ uptake develops in rats upon aging and show that in vivo administration of 1,25(OH)₂D₃ or its precursor to senescent rats restores the ability of the hormone to stimulate duodenal cell calcium influx through the cAMP messenger system. Received: 26 December 1997 / Accepted: 12 May 1998     

Improved renal transplant preservation using a modified intracellular flush solution (PB-2)

Summary A number of new intracellular renal flush solutions have been found to be more efficacious than Collins-2 (C-2) solution in extending organ viability during simple cold storage. However, the mechanism of action of these solutions remains poorly understood. To delineate better underlying intracellular mechanisms, we studied a modified, simple, hypothermic, intracellular (340 mOsm/kg) flush solution (PB-2). The development of PB-2 solution is based on the ability of some of its individual components to minimize ischemic adenine nucleotide (AN) catabolism and endothelial post reperfusion injury. Preliminary results in 10 canine autorenal transplants show a significant (P<0.02) improvement in renal recovery and viability (recipient posttransplant inulin clearance and survival) after 50 h of cold storage compared with 10 canine kidneys similarly preserved using conventional C-2 flush solution. High performance liquid chromotography (HPLC) studies show a significant (P<0.01) loss of AN using C-2, while PB-2 was associated with regeneration of AN within 45 min of reperfusion. Magnetic resonance spectroscopy using phosphorus 31 (³¹P-MRS) showed more high energy phosphorus metabolites (phosphomonoester and nicotinamide-adenine-dinucleotide phosphate: P<0.001) at 50 h cold storage using PB-2 compared with C-2. Electron micrographs (EM) revealed normal microcapillary morphology for the PB-2 group; however, moderate vascular red and white blood cell clumping was observed in the C-2 group. Characterization of the basic preservation mechanisms by HPLC, ³¹P-MRS, and EM studies indicates that PB-2 solution enhances renal preservation by diminution of both reperfusion injury and the loss of intracellular high energy metabolites that are necessary for viability.