A novel evaluation system to monitor bone formation and ß-tricalcium phosphate resorption in opening wedge high tibial osteotomy.

PURPOSE: The aim of this study was to establish an evaluation system to monitor bone formation and beta-tricalcium phosphate (TCP) resorption in opening high tibial osteotomy (HTO). METHODS: From 2003 to 2005, opening HTO was performed in 36 patients using a Puddu plate and ß-TCP blocks with 60 and 75 % porosity. Thirty-one patients were used for evaluation. All patients underwent CT examination at 2 weeks and 6 years. The CT image data were divided into three areas, and CT values of each area were analysed using the imaging software, Osirix. RESULTS: CT image analysis at 2 weeks showed that the mean CT-attenuation values (in Hounsfield units) of the implanted area with ß-TCP of 60 % porosity, the implanted area with ß-TCP of 75 % porosity, and cancellous bone were, 1,694.0 ± 94.2, 1,010.9 ± 81.1, and 178.0 ± 45.1, respectively. Six years after surgery, these values were 574.1 ± 273.5, 168.8 ± 75.1, and 174.9 ± 69.3, respectively. CONCLUSION: ß-TCP with 75 % porosity was completely resorbed and replaced by bone. ß-TCP with 60 % porosity was resorbed, but approximately 1/3 still remained even 6 years after surgery. The imaging software, Osirix, enabled scanning of the whole area to measure CT values. This system is the first to quantitatively evaluate ß-TCP resorption and bone formation in opening HTO. LEVEL OF EVIDENCE: Laboratory studies.

An Injectable Complex of ß-tricalcium Phosphate Granules, Hyaluronate, and rhFGF-2 on Repair of Long-bone Fractures with Large Fragments.

We evaluated the effects of an injectable complex of ß-tricalcium phosphate (ß-TCP) granules, hyaluronate, and recombinant human fibroblast growth factor-2 (rhFGF-2) on repair of unstable intertrochanteric fractures in elderly patients. Twenty-five patients (range, 76-91 years) having 31.A2 fractures (AO classification) were treated with injection of the complex followed by intramedullary nails. Bone regeneration and ß-TCP resorption, unions of intertrochanteric fractures and displaced lesser trochanters to the shaft, and varus deformity of the femoral neck were assessed by X-ray and CT scans. Fracture union occurred in all cases and union of the displaced lesser trochanter to the shaft was obtained in 24 cases by 12 weeks. It is of interest that ß-TCP granules were completely resorbed and marked new bone formation around the lesser trochanter was observed in all cases compared to cases not treated with the complex. Based on the results of intertrochanteric fractures, we applied this technique to two patients with subtrochanteric or humeral fractures in elderly patients, and obtained bone union. This complex is a paste-like material that is easy to handle, and it may be of considerable use in treatment of both unstable intertrochanteric fractures and other cortical bone defects with minimal surgical invasion.

Distinguishing adrenal adenomas from non-adenomas on dynamic enhanced CT: a comparison of 5 and 10 min delays after intravenous contrast medium injection.

AIM: To evaluate the usefulness of several parameters of 5 min compared to 10 min delayed contrast-enhanced CT in distinguishing adenomas from non-adenomas. MATERIALS AND METHODS: The study population consisted of 94 patients (52 men and 42 women; mean age 62 years) with 103 adrenal lesions (75 adenomas and 28 non-adenomas). In each patient, unenhanced CT was followed by early, 5 and 10 min enhanced CT. Diagnostic parameters included delayed enhanced attenuation at 5 and 10 min, washout attenuation (WO) at 5 and 10 min, absolute percentage washout (APW) at 5 and 10 min, and relative percentage washout (RPW) at 5 and 10 min. The accuracy of each parameter for diagnosing adenomas from non-adenomas was calculated using receiver operating characteristic (ROC) analysis. RESULTS: Upon comparison between 5 and 10 min delayed contrast-enhanced CT for differentiating total adenomas or lipid-poor adenomas from non-adenomas, there was no significant difference in the area under the binomial ROC curve (Az) values of delayed enhanced attenuation (total adenomas versus non-adenomas, p = 0.164; lipid-poor adenomas versus non-adenomas, p = 0.178), WO (total adenomas versus non-adenomas, p = 0.216; lipid-poor adenomas versus non-adenomas, p = 0.230), APW (total adenomas versus non-adenomas, p = 0.401; lipid-poor adenomas versus non-adenomas, p = 0.870), or RPW (total adenomas versus non-adenomas, p = 0.160; lipid-poor adenomas versus non-adenomas, p = 0.780). CONCLUSION: Five minute contrast-enhanced CT was as useful as 10 min contrast-enhanced CT for differentiation of adrenal adenomas from non-adenomas.

Use of an Injectable Complex of ß-Tricalcium Phosphate Granules, Hyaluronate, and Fibroblast Growth Factor-2 on Repair of Unstable Intertrochanteric Fractures.

We evaluated effects of an injectable complex of ß-tricalcium phosphate (ß-TCP) granules, hyaluronate, and recombinant human fibroblast growth factor-2 (rhFGF-2) on repair of unstable intertrochanteric fractures in elderly patients. Twenty-five patients (range, 76-91 years) having 31.A2 fractures (AO classification) were treated with injection of the complex followed by intramedullary nails. Bone regeneration and ß-TCP resorption, unions of intertrochanteric fractures and displaced lesser trochanters to the shaft, and varus deformity of the femoral neck were assessed by X-ray and CT scans. Fracture union occurred in all cases and union of the displaced lesser trochanter to the shaft was obtained in 24 cases by 12 weeks. It is of interest that ß-TCP granules were completely replaced by bone and marked new bone formation around the lesser trochanter was observed in all cases compared to cases not treated with the complex. This complex is a paste-like material that is easy to handle, and it may be of considerable use in treatment of both unstable intertrochanteric fractures and other cortical bone defects with minimal surgical invasion.

Long-term results of anterior cruciate ligament reconstruction using semitendinosus and gracilis tendons with Kennedy ligament augmentation device compared with patellar tendon autografts.

ACL reconstructions were postoperatively evaluated in 102 patients who underwent surgery using an arthroscopically assisted double-incision technique. Augmentation of the distally based semitendinosus and gracilis tendons was done either with the Kennedy ligament augmentation device (LAD) (Group 1 patients, mean follow-up of 108 months) or bone-patellar-tendon-bone graft (Group 2 patients, mean follow-up of 109 months). At follow-up, serial KT-1000 measurements showed < or = 5 mm side-to-side differences in 70% of Group 1 and 90% of Group 2 patients. No differences were found among patients regarding postreconstruction complications such as loss in motion range, patellofemoral crepitus, osteoarthritis, and muscle disfunction.

Comparison of biochemical characteristics of cultured fibrochondrocytes isolated from the inner and outer regions of human meniscus.

In order to evaluate the ability of fibrochondrocytes to synthesize collagen and proteoglycan, human medial meniscal cells were cultured in a monolayer. Meniscal cells were prepared from the two regions (outer 1/3 and inner 2/3) in consideration of the difference in vascular supply, and articular chondrocytes were also obtained from the same knee joint. Regarding total collagen synthesis, regional differences were not found, but age-related differences were found in human medial meniscus. In contrast, proteoglycan synthesis revealed significant regional differences; meniscal cells from the inner 2/3 synthesized a greater amount of proteoglycan. After long-term monolayer culturing, proteoglycan synthesis by meniscal cells decreased in a time-dependent manner, and morphological changes to fibroblast-like cells were found. In the presence of transforming growth factor (TGF)-beta, proteoglycan synthesis increased in a dose-dependent manner. These findings suggest that the inner regions of the human meniscus contain cells with a chondrocytic phenotype.

Human c-Jun N-terminal kinase expression and activation in the nervous system.

Differential expression and localization of c-Jun N-terminal kinases (JNKs) in the human brain may reflect transduction of a variety of extracellular stimuli to selective cellular responses. Of the three JNKs, JNK1 and 2 are widely distributed in tissues and JNK3 is predominantly restricted to brain where it is expressed in neurons. Although there is considerable molecular conservation among all three JNKs, we distinguished expression of each by in situ hybridization, immunoblot analysis with a panel of antibodies, and stress-activation using c-Jun as substrate. In the human central nervous system (CNS), there are at least 10 isoforms: JNK3alpha1 and JNK1alpha1 were the major JNK isoforms expressed; JNK2 was not detected. On immunoblots of brain homogenates, antibody selectivity identified JNK3alpha1 as a 45-kDa protein, JNK1alpha1, a slightly lower band at 44 kDa, and a 50-kDa band of unknown specificity. Recombinant human JNK3alpha1, transfected either into CHO, COS-1, or Neuro2A (N2A) cells, was strongly expressed as a 45-kDa protein in each. Transfected JNK3alpha1, and endogenous JNK1, each immunoprecipitated from N2A cells, phosphorylated recombinant forms of human c-Jun. Kinase activity of each JNK was modestly stimulated in N2A cells by anisomycin but not by ceramide, UV irradiation, or heat shock. Endogenous JNK activation, especially at a low level, may reflect a chronic and cumulative stress process that contributes to hyperphosphorylation of cytoskeletal proteins such as those found in Alzheimer's disease (AD), and ultimately, induction of apoptosis.

[Proteoglycan and collagen synthesis of cultured fibrochondrocytes from the human knee joint meniscus].

Fibrochondrocytes isolated from different parts of the human knee joint meniscus were cultured, and their morphology and ability to synthesize matrix components were investigated. The mid-portion of the medial meniscus was divided into two parts (the outer one-third and the inner two-thirds), and cells were obtained for culture by pronase and collagenase digestion. The appearance of the fibrochondrocytes was different in monolayer culture from that of both articular chondrocytes and fibroblasts. However, there was no difference between the cells obtained from the two different parts of the meniscus. Although proteoglycan synthesis by cells from the inner two-thirds was higher than that by cells from the outer one-third of the meniscus, there was no difference in the collagen synthesis. Fibrochondrocytes from the meniscus were found to synthesize cartilage-like proteoglycan. Proteoglycan synthesis by meniscus cells and articular chondrocytes was stimulated by TGF-beta 1 in a dosage-dependent manner. These findings indicated that fibrochondrocytes possessed similar characteristics to articular chondrocytes in terms of proteoglycan metabolism.

The toxic effect of Alzheimer amyloid protein precursor overexpressed in the neuroblastoma cell line NB-1 on neurite outgrowth.

The neuroblastoma cell line NB-1 is induced to start neurite outgrowth by dibutyryl cyclic AMP (Bt2cAMP). To study the function of Alzheimer amyloid protein precursor (APP), a stable cell line overexpressing APP was established by cDNA transfection. The APP cDNA was driven by the cytomegalovirus early gene promoter. The transformant underwent degeneration as well as neurite outgrowth in the presence of Bt2cAMP, which also increased the amount of APP mRNA and protein. The expressed APP was mainly processed in the secretory pathway and few amyloidogenic fragments were observed. Hence, not only the beta/A4 protein but also the overexpressed APP itself might be neurotoxic.

Monoamine oxidase inhibitors prevent striatal neuronal necrosis induced by transient forebrain ischemia.

Seven days after 30 min of ischemia, neuronal necrosis was observed in the striatum. Pretreatment with type A monoamine oxidase (MAO-A) inhibitors, clorgyline and RS-8359 ((+)-4-(4-cyanoanilino)-7-hydroxycyclopenta (3,2-e) pyrimidine) decreased significantly the number of necrotic neurons and inhibited changes in the dopamine metabolite contents during and after transient ischemia. An MAO-B inhibitor, deprenyl also decreased the neuronal necrosis, but it inhibited only the changes in 3,4-dihydroxyphenylacetic acid (DOPAC) content after reperfusion. The results suggest that the activation of dopamine metabolism after transient ischemia was mainly mediated by MAO-A and partly by MAO-B and suggest a possible role of dopamine deamination by MAO in the development of ischemic neuronal necrosis.

Output, tissue levels, and synthesis of acetylcholine during and after transient forebrain ischemia in the rat.

Biochemical changes in the rat brain cholinergic system during and after 60 min of ischemia were studied using a four-vessel occlusion model. Extracellular acetylcholine (ACh) concentrations in the unanesthetized rat hippocampus markedly increased during ischemia and reached a peak (about 13.5 times baseline levels) at 5-10 min after the onset of ischemia. At 2-5 h after reperfusion, extracellular ACh concentrations were reduced to 64-72% of the levels of controls. ACh levels in the hippocampus, striatum, and cortex decreased significantly during ischemia and exceeded their control values just after reperfusion. A significant increase in hippocampal ACh level after 2 days of reperfusion and a decrease in [14C]ACh synthesis from [14C]glucose in hippocampal slices excised at 2 days after reperfusion were observed. The extracellular concentrations and tissue levels of choline markedly increased after ischemia. These results show that ACh is markedly released into the extracellular space in the hippocampus during ischemia, and they suggest that ACh synthesis is activated just after reperfusion and that cholinergic activity is reduced after 2-48 h of reperfusion in the hippocampus.

Deamination of norepinephrine, dopamine, and serotonin by type A monoamine oxidase in discrete regions of the rat brain and inhibition by RS-8359.

Levels of monoamines and their metabolites were determined in the cortex, hippocampus, and striatum of rats killed by microwave irradiation. Moclobemide (20 mg/kg, p.o.) and clorgyline (10 mg/kg, p.o.), type A monoamine oxidase (MAO-A) inhibitors, increased the levels of normetanephrine (NM) and 3-methoxytyramine (3MT) and decreased those of 3,4-dihydroxyphenylacetic acid (DOPAC), homovanillic acid (HVA), and 5-hydroxyindoleacetic acid (5HIAA) in almost all three regions. Deprenyl (10 mg/kg, p.o.), a type B monoamine oxidase inhibitor, however, little affected monoamine and metabolite levels in all regions. The maximum effects of RS-8359 (10 mg/kg, p.o.) were obtained at 2 to 6 hr after administration, when the levels of norepinephrine (NE), NM, 3MT, and serotonin (5HT) in all regions and dopamine (DA) in the striatum increased, while DOPAC and HVA levels decreased. The levels of monoamines and metabolites had returned to normal by 20 hr after administration. Dose-dependency of the effects of RS-8359 on monoamine metabolites was observed at doses up to 30 mg/kg (p.o.) at 1 and 6 hr after administration. In conclusion, NE, DA, and 5HT are exclusively or preferentially deaminated by MAO-A in the cortex, hippocampus, and striatum of rats, and RS-8359 exhibits a reversible MAO-A inhibitory action in all three regions tested in vivo.

Participation of type A monoamine oxidase in the activated deamination of brain monoamines shortly after reperfusion in rats.

Changes in monoamine levels during and after ischemia and effects of RS-8359, a type A monoamine oxidase (MAO-A) inhibitor, were studied in the cerebral cortex, hippocampus, and striatum of rats killed by microwave irradiation. The patterns of the changes in norepinephrine (NE), dopamine (DA), and serotonin (5HT) levels were similar during ischemia: All these monoamines decreased in all three regions. After reperfusion, however, the three monoamines showed different patterns of changes: NE, except in the striatum, decreased further; DA increased over the controls; 5HT remained suppressed in all three regions. With regard to the metabolites of the monoamines, the changes during and after reperfusion were almost similar in all regions: O-methylated metabolites, normetanephrine and 3-methoxytyramine, markedly increased during ischemia; After reperfusion, the elevated levels of normetanephrine and 3-methoxytyramine returned to normal, while deaminated metabolites, 3,4-dihydroxyphenylacetic acid (DOPAC) and 5-hydroxyindoleacetic acid, homovanillic acid (HVA), and 3-methoxy-4-hydroxy-phenylethyleneglycol clearly increased. RS-8359 pretreatment (30 mg/kg, p.o.) at an hour prior to ischemia elevated the levels of NE in the cortex and hippocampus during ischemia and inhibited the increases in DOPAC and HVA levels and the decrease in 3MT levels at 30 min after reperfusion. These results suggest that deamination of NE, DA, and 5HT is activated by the increases in the substrates for MAO in all three regions, except the noradrenergic system in the striatum, and that MAO-A participates in the activated deamination after reperfusion.

[3H]thymidine autoradiographic and alkaline phosphatase histochemical studies of intestinal metaplasia of the human stomach.

The relationship between cell proliferation and enzyme activity in intestinal metaplasia of the human stomach was studied using a combined method of [3H]thymidine autoradiography and alkaline phosphatase histochemistry on the same section. Three types of intestinal metaplasia were observed depending on variations in both enzymatic activity and isotope labelling. One type shows alkaline phosphatase-positive cells along the entire length of the glands with [3H]thymidine-labelled cells localized only at the bottom of the glands, resembling the duodenum. In another type of intestinal metaplasia, alkaline phosphatase-positive cells are present on the surface and/or upper half of the glands with mitotically active cells occupying the lower part of the glands. The third variety of intestinal metaplasia is characterized by the absence of alkaline-phosphatase activity and [3H]thymidine-labelled cells present in an extended zone in the lower half of the glands. Differences in labelling patterns of [3H]thymidine and the activity of marker enzyme in various types of intestinal metaplasia seem to reflect variations in cell differentiation during intestinalization of gastric mucosa.

Cytochemical localization of alkaline phosphatase in intestinal metaplasia of the human stomach.

Alkaline phosphatase in the brush border of areas of intestinal metaplasia of human stomach was studied cytochemically. All absorptive cells in the upper part of the villi of the duodenum had strong alkaline phosphatase activity but, in areas of intestinal metaplasia, the metaplastic glands consisted of alkaline phosphatase-positive and negative absorptive cells. Alkaline phosphatase activity was found in tall dense microvilli of absorptive cells in areas of intestinal metaplasia and in the duodenum. However, in some areas of metaplastic epithelium the activity was very weak in some tall dense microvilli of absorptive cells but strong in those of neighbouring absorptive cells. No alkaline phosphatase activity was found in short sparse microvilli of absorptive cells in areas of intestinal metaplasia. The difference in alkaline phosphatase activity in microvilli of different cells in areas of intestinal metaplasia, which is not seen in the duodenum, indicates abnormal morphological and enzymatic differentiation intestinal metaplasia.