Expression of transglutaminase 2 does not differentiate focal myositis from generalized inflammatory myopathies

Objectives –  Idiopathic inflammatory myopathies (IIM), including dermatomyositis (DM), polymyositis (PM), sporadic inclusion-body myositis (s-IBM) and focal myositis (FM) are a heterogeneous group of autoimmune disorders of skeletal muscle. An increased transglutaminase 2 (TG2) expression has been found in DM, PM and s-IBM. The aim of our study was to investigate TG2 expression in FM in comparison with other IIM.
Materials and methods –  We re-examined tissue material we have gathered in the course of our previous studies on IIM, investigating muscle expression of TG2 in patients with FM in comparison with DM, PM and s-IBM using immunocytochemistry and real-time RT-PCR.
Results –  Immunocytochemistry revealed an increased TG2 signal in endomysial vessels, in atrophic and degenerating/regenerating muscle fibres in PM, DM, s-IBM and FM; in s-IBM, some vacuoles were immunostained too. Real-time RT-PCR study confirmed a significantly increased expression of TG2 in all IIM muscles examined.
Conclusions –  Our study demonstrates the presence of TG2 in FM muscles. The study suggests that TG2 expression does not represent a distinctive marker to differentiate FM from generalized IIM. TG2 over-expression in inflamed skeletal muscle does not seem have a pathogenetic role in such a disease, but it could represent a way to contain the inflammatory process.     

Distal‐symmetric focal inflammatory myopathy distinct from focal myositis and polymyositis

We describe an inflammatory myopathy that was symmetrically restricted to both gastrocnemius muscles in a young man. Histopathological findings were typical for polymyositis, but there were neither signs of generalization nor muscle weakness and wasting. This condition was highly sensitive to steroids and has been kept in remission for more than a year using azathioprine. Our findings add another entity to the spectrum of spatially restricted inflammatory myopathies. Muscle Nerve 40: 309–312, 2009     

HLA typing in focal myositis

It is still controversial if idiopathic focal myositis is a part of systemic polymyositis. We present here four patients, including identical twins, with focal myositis accompanied by the same HLA typings. Gradually developing unilateral calf muscle pain was an initial symptom in all patients. Neither muscular weakness nor creatine kinase (CK) elevation was observed, while minimal inflammatory findings such as erythrocyte sedimentation rate (ESR) increase appeared in serum. Magnetic resonance imaging (MRI) revealed localized abnormalities of calf muscles. Biopsy specimen was characterized by perimysial and endomysial inflammatory infiltration consisted of T cells and macrophages and rare necrotic fibers. Corticosteroid administrations ameliorated their symptoms and signs, though recurrence occurred along with decreasing doses. HLA typings common to all patients were A2, B62, Cw3, and DQ3, whereas HLA-D DNA typings were DQB1 *0303 for two patients, and DQB1*0302 for three patients. These findings suggest that at least some focal myositis may be a new disease unit, with a common genetic background but not a part of systemic polymyositis.     

Camptocormia in Parkinson's disease mimicked by focal myositis of the paraspinal muscles.

We report on a 63-year-old man with idiopathic Parkinson's disease who developed kyphosis and a severe forward flexion of the thoracolumbar spine. A typical feature was an increase during walking or standing and it completely disappeared in the supine position, mimicking the clinical phenomenon of camptocormia (bent spine). In addition to the abnormal posture, a weakness of the erector spinal muscles, local pain, reddening, and elevated temperature of the paraspinal muscles were evident. Creatine kinase was initially elevated, electromyography showed spontaneous activity and a myopathic pattern. Magnetic resonance imaging and bioptic examinations revealed a focal myositis of the paraspinal muscles. This case indicates that camptocormia can be mimicked by focal myositis of paraspinal muscles and must be included in the differential diagnosis, especially when additional symptoms as inflammatory signs or weakness are present.     

The value and correlation between PRL‐3 expression and matrix metalloproteinase activity and expression in human gliomas

Local invasion of tumor cells is characteristic of most human glioma invasions. It is associated with increased motility and a potential to degrade the extracellular matrix. Matrix metalloproteinases (MMPs) have been proved to be a main process in local invasion of brain tumor. PRL‐3 is a new protein tyrosine phosphatase which would also degrade the extracellular matrix and has been proved to be expressed in liver metastases derived from colorectal cancer. In this study, we sought to investigate the expression of PRL‐3 in glioma tissues and investigate the relationship between MMPs (MMP2, MMP9, membrane‐type matrix metalloproteinase 1 [MT1‐MMP]) activity and expression in gliomas. The modifications of in situ hybridization of mRNA phosphatase of regenerating liver‐3 (PRL‐3) methods are preformed in the study of paraffin‐embedded slides. The immunohistochemistry and gelatin zymography are used to detect the expression of PRL‐3 and activity of MMPs. The results show that PRL‐3 mRNA and antibody of PRL‐3 are detected in glioma tissues mainly in grades IV and III, only a little in grade II, but not in normal brain tissue and glioma grade I. MMP2 and MMP9 are observed mainly in glioma tissues of grades IV and III in activity and expression. MT1‐MMP protein is located in glioma tissues and vessel endothelial cells. This is the first report of detecting PRL‐3 expression in gliomas, especially in grades III and IV, which may play an important role in progression of gliomas. PRL‐3, MMP2 and MT1‐MMP cooperatively contribute to gliomas invasion. Intermediate MMP2 (MT1‐MMP, TIMP‐2, MMP2 trimeric complex) is detected in high grades of glioma tissues by gelatin zymography and may be a marker indicating latent malignance of gliomas.     

rt-PA应用后MMP-2、MMP-9表达的改变及Neuroserpin的影响

目的 观察重组组织型纤溶酶原激活剂(rt-PA)对血管再通后基质金属蛋白酶-2(MMP-2)、MMP-9表达的影响以及神经源性丝氨酸蛋白酶抑制剂(neuroserpin,NSP)的干预作用。方法 应用易卒中型肾血管性高血压大鼠复制大脑中动脉缺血模型,缺血3 h后再灌注并静脉注射rt-PA,于预组在应用rt-PA前脑内注射NSP,1天后处死,常规病理检查,并应用免疫组织化学和原位杂交的方法观察MMP-2、MMP-9在脑组织的表达。结果 缺血再灌注后MMP-2、MMP-9表达均升高;应用rt-PA后可见缺血再灌注区有灶性出血及红细胞漏出,同时使MMP-9进一步升高,但对MMP-2影响不大;应用rt-PA的同时使用NSP可以减轻缺血损伤,减少出血的发生,并使升高的MMP-9减少至接近正常水平,但NSP可以使MMP-2表达略有升高。结论 rt-PA溶栓后出血转化的发生可能与MMP-9表达增加有关,溶栓时联合应用NSP可能通过降低rt-PA所致的MMP-9表达上调而减轻溶栓治疗的出血并发症。     

Focal myositis associated with S-1 radiculopathy: report of two cases

Two cases are described of pseudotumoral calf hypertrophy after laminectomy for a compressive S-1 radiculopathy. The serum creatine kinase (CK) level was normal or mildly elevated. T2-weighted magnetic resonance imaging (MRI) showed calf enlargement, with an increased signal of the medial head of the gastrocnemius muscle. Electromyography revealed fibrillation potentials and positive sharp waves, but no complex repetitive discharges in the affected gastrocnemius muscle, with motor unit potentials having mixed neurogenic and myopathic features. Muscle biopsy revealed a focal myositis associated with some features of denervation. A brief course of corticosteroids was followed by remission clinically and improvement in the MRI findings.     

Granulomatous myositis: a clinical study of thirteen cases

Granulomatous myositis (GM) is a rare condition that has generally been described in association with sarcoidosis. In the absence of sarcoidosis or other underlying disease, a diagnosis of isolated GM is considered. Only one study has focused on the clinical difference between isolated GM and sarcoid myopathy (SM). We report 13 cases of symptomatic GM; 8 had sarcoidosis. All patients with sarcoidosis had predominantly proximal, symmetrical lower-limb weakness, and 3 subsequently developed upper-limb or distal involvement. Three of the five patients with isolated GM had predominantly distal muscle involvement, and two had dysphagia. Corticosteroid treatment was followed by prolonged improvement in only one patient with sarcoidosis. One patient had acute sarcoid myositis and benefited from methotrexate; other immunosuppressants and etanercept proved ineffective in chronic sarcoid myopathy. Three of the five patients with isolated GM responded to corticosteroid treatment. When last examined, three patients with sarcoidosis had severe disability, whereas patients with isolated GM showed milder weakness. Thus, SM was frequently associated with severe disability and rarely improved after corticosteroid treatment, whereas most patients with isolated GM improved.     

Camptocormia associated with focal myositis in multiple-system atrophy.

Camptocormia (CC) or pronounced forward flexion of the trunk is a common symptom of Parkinson's disease. We describe 2 patients with probable, respectively possible multiple-system atrophy and CC. Magnetic resonance imaging of the erector trunci showed focal patchy hyperintensities with gadolinium enhancement and muscle biopsy was indicative of variably pronounced focal myositis. CC was progressive and the major handicap for both patients after 1 and 1.5 years of follow-up, respectively. The therapeutic response was poor. Similarities with the dropped-head syndrome suggest that the muscle pathology may be either the primary cause of CC, a focal reaction to the CC posture, or a coincident syndrome of old age.     

Adhesion molecule expression in experimental myositis

Experimental allergic myositis (EAM) in Lewis rats, induced with partially purified myosin, is regarded as a model of human polymyositis. To clarify the role of adhesion molecules in the pathogenesis of EAM in Lewis rats, we investigated intramysial expressions of the intercellular adhesion molecule (ICAM)-1 and vascular cell adhesion molecule (VCAM)-1, and the serum level of soluble ICAM-1 in EAM rats. All the EAM rat muscles had scattered inflammatory foci, as well as cell infiltration and necrosis, by week 4 after the initial immunization (i.e., day 0 after the last immunization). As compared with the control muscles, ICAM-1 and VCAM-1 were strongly expressed immunohistochemically in the endothelium of vessels in the endomysium and perimysium, and to lesser extents in the inflammatory infiltrates and on the sarcolemma of nonnecrotic muscle fibers adjacent to the inflammatory infiltrates or invaded muscle fibers. ICAM-1 in the muscle extracts and sera from EAM rats increased on each test day, as compared with extracts from the normal controls. The values peaked on day 0 after the last immunization, then gradually decreased with time. ICAM-1 elevations in the muscle extracts were correlated with the percent of sections that had inflammatory lesions (P = 0.032) and the histological scores (P = 0.005) on day 0, whereas there was no significance on days 3 and 7. These findings suggest that the adhesion molecules ICAM-1 and VCAM-1 increase in the early stage of EAM, and function in the initiation of the inflammatory process of myositis.     

A novel method to establish microglia-free astrocyte cultures: comparison of matrix metalloproteinase expression profiles in pure cultures of astrocytes and microglia

Increased matrix metalloproteinase (MMP) proteolytic activity contributes to the pathogenesis of many neuroinflammatory and neurodegenerative conditions in the CNS. To fully understand this process, it is important to define the MMP expression profile of specific cell types, including the CNS-resident cells astrocytes and microglia. While previous studies have characterized astrocyte MMP expression by using mixed glial cultures, these results are likely complicated by the presence of contaminating microglia within these cultures. In the current study, we sought to clarify this complexity, by taking a novel approach to prepare pure astrocyte cultures entirely devoid of microglia, by promoting neural stem cell (NSC) differentiation into astrocytes. The MMP expression profile of mixed glial cultures, neurosphere-derived astrocytes, and pure microglia was characterized by RNase protection assay. This revealed that MMP gene expression is largely cell-type specific. Astrocytes constitutively expressed MMP-11, MMP-14, and MMP-2 and showed induction of MMP-3 in response to IL-1beta but did not respond to lipopolysaccharide (LPS). In contrast, microglia constitutively expressed high levels of MMP-12 and showed strong induction of MMP-9 and MMP-14 in response to LPS. Gelatin zymography confirmed that LPS and TNF-alpha induced strong expression of MMP-9 in microglia but not astrocytes. In summary, these studies demonstrate that neurosphere-derived astrocytes represent an attractive alternative system in which to study astrocyte behavior in vitro. Using this system, we have shown that astrocytes and microglia express distinct sets of MMP genes and that microglia, not astrocytes, are the major source of MMP-9 in response to LPS or TNF-alpha.     

Comparison of matrix metalloproteinase activation after focal cortical ischemia in young adult and aged mice

Matrix metalloproteinase (MMP) activity is implicated in the degradation of the extracellular matrix during cerebral ischemia. Although many studies have demonstrated spatiotemporal patterns of activation of gelatinases (MMP-9 and MMP-2) after ischemic stroke in young adult rodents, no data exist on MMP activity in old brains. In this study, we investigated the gelatinolytic activity in young adult (3-month-old) and aged (1-year-old) mice subjected to photothrombotic stroke. Using in situ zymography and gel zymography, we found that the basal gelatinolytic activity in the intact cerebral cortex was similar at both investigated ages. Similarly, after photothrombosis, the increased gelatinolytic response up to 7 days poststroke was the same in young and aged brains. At both ages, early activation of gelatinolysis in the ischemic core and the perilesional area was present in neuronal nuclei as revealed by colocalization of gelatinolytic product with NeuN immunostaining and DAPI. Additionally, application of specific antibodies against MMP-9 and MMP-2 revealed the increase in MMP-9 immunoreactivity in cell nuclei as early as 4 hr poststroke. No differences between young and aged mice were observed concerning the level and localization of MMP-9 immunoreactivity. The lack of age-related differences in the degree and pattern of activation of gelatinolysis after focal stroke and the lack of correspondence between the results of in situ and gel zymography suggest that extracellular proteolysis is not directly responsible for the more severe outcome of ischemic stroke in aged subjects.     

Alterations in matrix metalloproteinase-9 levels and tissue inhibitor of matrix metalloproteinases-1 expression in a transforming growth factor-beta transgenic model of hydrocephalus

The development of spontaneous hydrocephalus in mouse models resulting from the overexpression of transforming growth factor-beta (TGFbeta-1) has been previously described, although the mechanism by which this occurs remains obscure. It has been previously demonstrated that increased expression of TGFbeta has consequences for the levels of matrix metalloproteinases (MMPs) and their specific inhibitors (tissue inhibitors of MMPs, or TIMPs). These remodeling proteins play an important role in extracellular matrix (ECM) maintenance through degradation and deposition of ECM components. The present study investigated the relationship between elevated levels of TGFbeta-1, the ECM modulators TIMP-1 and MMP-9, and development of hydrocephalus in the neonatal mouse. In newborn pups, TIMP-1 mRNA levels were equal between animals expressing the TGFbeta-1 transgene and littermates without the transgene. However, immunohistochemistry of littermate pups shows that the distribution of TIMP-1 was changed from homogeneous with large punctate concentrations of signal to uniform, dense staining in hydrocephalic animals carrying the TGFbeta-1 transgene. The mRNA levels of MMP-9 were decreased in the transgenic animals, as were the activity levels MMP-9. These results suggest that the remodeling protein MMP-9 and its specific inhibitor, TIMP-1, may contribute to the spontaneous development of hydrocephalus in this transgenic model by altering the ECM environment.     

Expression of matrix metalloproteinases in the neurogenic niche of the adult monkey hippocampus after ischemia

Matrix metalloproteinases (MMPs), zinc-dependent endopeptidases capable of remodeling extracellular matrix and regulating cellular signals, have been implicated in various neurological functions and diseases. However, the role of MMPs in the adult neurogenesis still remains to be clarified, particularly in the primate. Here, we studied differential expression of MMP9/2 in the neurogenic niche of the hippocampal dentate gyrus (DG) after transient global brain ischemia in young adult macaque monkeys. Zymography demonstrated biphasic upregulation of MMP9 in acute (Days 1-3) and delayed (Days 7-15) phases of postischemic reaction, whereas the level of MMP2 was elevated only in the delayed phase. Immunofluorescence histochemistry showed that MMP9 and MMP2 colabeled with markers of endothelial cells, astrocytes, and newborn neurons in the subgranular zone (SGZ) of the DG, and also that the percentage of coexpression significantly increased in the delayed postischemic phase, as compared with controls. However, colabeling with different cell selective markers reached its peak at different time points, i.e., with endothelial cells on Day 7, whereas with astrocytes and newborn neurons on Day 15, respectively. MMPs were localized both in the perikarya and dendrites in the newborn neurons. In conclusion, MMP9/2 expression was regulated in a cell- and time-specific manner in hippocampal neurogenic niche of adult primates.     

High-dose immunoglobulin therapy in sporadic inclusion body myositis: a double-blind, placebo-controlled study

Sporadic inclusion body myositis (s-IBM) is an acquired inflammatory muscle disease of unknown cause. In general, s-IBM presents with slowly progressive, asymmetric weakness, and atrophy of skeletal muscle. There is a mild transitory or nil responsiveness to standard immunosuppressive treatment. A controlled cross-over study of 22 s-IBM patients over 3 months showed a partial improvement in those treated with high-dose intravenous immunoglobulin therapy (IVIG) versus placebo. The present study included 22 patients aged 32-75 years and with a mean duration of disease of 5.2+/-3.6 years. They were randomized by a double-blind, placebo-controlled, cross-over design to monthly infusions of 2 g/kg bodyweight IVIG or to placebo for 6 months each, followed by the alternative treatment. After 6 and 12 months the response to treatment was evaluated, using a modified Medical Research Council scale, Neuromuscular Symptom Score (NSS), the patient's own assessment of improvement, arm outstretched time, and electromyography. No serious side effects were seen, in particular no viral infection and no major cardiac or neurological complications. Overall there was no progression of the disease in 90% of patients, unlike that which might have been expected in untreated patients. A mild and significant improvement (11%) in clinical symptoms was found using NSS, but not with other test procedures. There was a trend to mild improvement in treated patients when using other tests. Individual responses to treatment was heterogeneous. The validity of this study may be reduced by mismatch of groups with regard to age at onset and variability in disease expression. The findings of this study largely confirm those of a previous IVIG trial. Treatment with IVIG may be mildly effective in s-IBM by preventing disease progression or inducing mild improvement. Long-term studies are needed to evaluate further the benefit of IVIG therapy in s-IBM.     

基质金属蛋白酶9的表达在慢性酒精中毒性肌损伤中的作用

目的 探讨基质金属蛋白酶9(MMP-9)在慢性酒精中毒性肌损伤(chronic alcoholic muscle injury,CAMI)发病过程中的作用.方法 60只雄性SD大鼠随机分为对照组(A组,15只,生理盐水灌胃)和实验组(B组,45只,灌酒),建立CAMI模型.将A组再随机分为A1～3组(分别于生理盐水灌胃第2周、第6周和第12周末处死,每组5只);B组再随机分为B1～3组(分别于灌酒第2周、第6周和第12周末处死,每组15只).光镜下观察大鼠骨骼肌病理学改变;逆转录-聚合酶链反应检测大鼠骨骼肌MMP-9的mRNA表达的动态改变;Western blot检测大鼠骨骼肌MMP-9蛋白的表达变化;明胶酶谱法检测大鼠骨骼肌MMP-9的酶活性变化.结果 组织化学结果显示,B组大鼠跖肌Ⅱ型肌纤维萎缩,间质炎性细胞浸润和纤维增生,其中I型胶原增生明显.A1组与B1组大鼠骨骼肌MMP-9 mRNA的表达甚少,Western blot和酶谱法检测仅显示MMP-9前体型.B2组大鼠跖肌MMP-9 mRNA表达(0.0579±0.0046)较A2组(0.0198±0.0023)增高(t=-9.151,P<0.01),B3组大鼠跖肌和腓肠肌MMP-9 mRNA的表达(0.1417±0.0116、0.0599±0.0057)均较A3组(0.0249±0.0049、0.0237±0.0021)增高(t=-18.095,P<0.01;t=-5.933,P<0.05).同时,Western blot和酶谱法也检测出B3组大鼠腓肠肌和跖肌活性型MMP-9的表达量及酶活性进一步增强,与B2组相比差异有统计学意义.结论 MMP-9的表达和活性增高可能参与了骨骼肌间质纤维增生,促进了CAMI.     

Celecoxib对糖尿病大鼠缺血再灌注脑组织ICAM-1和MMP9表达及神经功能的影响

目的探讨celecoxib(塞来昔布)对糖尿病大鼠脑缺血再灌注后脑组织中ICAM-1和MMP9表达及神经功能的影响。方法将SD大鼠分为假手术组(S组)、脑缺血再灌注生理盐水组(NS组)、celecoxib低剂量组(LCIR组)和高剂量组(HCIR组)。采用腹腔注射链脲佐菌素(STZ)和线栓法制作糖尿病大鼠大脑中动脉缺血再灌注模型。分别于缺血后30min给予生理盐水或celecoxib溶液灌胃,再灌注后6、12、24、48h将大鼠断头取脑,免疫组化法检测脑组织中ICAM-1和MMP9的表达,并对大鼠进行神经功能缺损评分和死亡率的比较。结果 NS组、LCIR组、HCIR组大鼠神经功能缺损评分有显著差异(P<0.05);NS组、LCIR组、HCIR组MMP9及ICAM-1阳性血管主要表达于缺血周边区,较S组表达明显增强(P<0.05),LCIR组、HCIR组MMP9及ICAM-1阳性血管数较NS组减少(P<0.05),LCIR组、HCIR组之间差异不明显(P>0.05),各组不同时间点之间MMP9及ICAM-1阳性血管数均有明显差异(P<0.05)。经Celecoxib干预后大鼠死亡率明显下降。结论 celecoxib可能通过抑制脑组织中MMP9和ICAM-1的表达而减轻糖尿病大鼠脑缺血-再灌注后神经功能的损伤而降低实验大鼠的死亡。     

Developmental regulation of membrane type-5 matrix metalloproteinase (MT5-MMP) expression in the rat nervous system

An intricate balance between extracellular matrix (ECM) synthesis and degradation must be maintained during developmental tissue remodeling. Matrix metalloproteinases (MMPs) are the main mediators of ECM degradation. A subset of MMPs, referred to as membrane-type MMPs, contains a transmembrane domain that restricts protease activity at the cell surface. Membrane type-5 MMP is predominantly expressed in the brain. The present report is the first to demonstrate the temporal regulation and spatial distribution of MT5-MMP mRNA during nervous system development.     

Cerebral embolization associated with parenchymal seeding of the left atrial myxoma: Potential role of interleukin‐6 and matrix metalloproteinases

Systemic embolization has been reported in up to 40% of patients with left atrial myxoma, half of them with cerebral involvement. However, development of intracerebral embolization associated with parenchymal seeding of the myxoma emboli is an extremely rare complication, with only 36 histologically diagnosed cases reported in the published literature. We describe a 69‐year‐old woman who arrived at the emergency service with hemiparesis associated with drug‐resistant epilepsy and a medical history of resection of a left atrial myxoma 10 months previously. Cranial computed tomography revealed multiple large lesions of heterogeneous density and cystic components in the occipital lobes and posterior fossa parenchyma. Histopathological analyses after stereotactic biopsy of the occipital lesion revealed infiltrative myxoma cells with benign histological findings and uniform expression of calretinin similar to that of the primary cardiac myxoma. Additional immunohistochemical studies confirmed brain parenchymal seeding of the myxoma cells with strong expression of interleukin‐6 (IL‐6) and focal expression of matrix metalloproteinases‐2 (MMP‐2). Here, we discuss the clinicopathological features of intracerebral embolization of left atrial myxomas associated with progressive parenchymal seeding of the tumor emboli and the potential pathogenic role of IL‐6 and MMPs.