Targeting ATR in vivo using the novel inhibitor VE-822 results in selective sensitization of pancreatic tumors to radiation.

Combined radiochemotherapy is the currently used therapy for locally advanced pancreatic ductal adenocarcinoma (PDAC), but normal tissue toxicity limits its application. Here we test the hypothesis that inhibition of ATR (ATM-Rad3-related) could increase the sensitivity of the cancer cells to radiation or chemotherapy without affecting normal cells. We tested VE-822, an ATR inhibitor, for in vitro and in vivo radiosensitization. Chk1 phosphorylation was used to indicate ATR activity, γH2AX and 53BP1 foci as evidence of DNA damage and Rad51 foci for homologous recombination activity. Sensitivity to radiation (XRT) and gemcitabine was measured with clonogenic assays in vitro and tumor growth delay in vivo. Murine intestinal damage was evaluated after abdominal XRT. VE-822 inhibited ATR in vitro and in vivo. VE-822 decreased maintenance of cell-cycle checkpoints, increased persistent DNA damage and decreased homologous recombination in irradiated cancer cells. VE-822 decreased survival of pancreatic cancer cells but not normal cells in response to XRT or gemcitabine. VE-822 markedly prolonged growth delay of pancreatic cancer xenografts after XRT and gemcitabine-based chemoradiation without augmenting normal cell or tissue toxicity. These findings support ATR inhibition as a promising new approach to improve the therapeutic ration of radiochemotherapy for patients with PDAC.

A quantitative analysis of lymphatic vessels in human breast cancer, based on LYVE-1 immunoreactivity.

This study was undertaken to determine the highly sensitive method for detecting tumour lymphatic vessels in all the fields of each slide (LV), lymphatic microvessel density (LMVD) and lymphatic vessel invasion (LVI) and to compare them with other prognostic parameters using immunohistochemical staining with polyclonal (PCAB) and monoclonal antibodies (MCAB) to the lymphatic vessel endothelial hyaluronan receptor-1 (LYVE-1), and the pan-endothelial marker factor VIII in a series of 67 human breast cancers. In all LYVE-1-stained sections, LV (some of which contained red blood cells) were frequently found localised in extralobular stroma, dermis, connective tissue stroma and adjacent to artery and vein, but were rare within the intralobular stroma or the tumour body (3/67 cases) or areas of widespread invasion. In contrast small blood vessels were observed in intra- and extralobular stroma in the factor VIII-stained sections. Quantitation of vessel numbers revealed that LYVE-1/PCAB detected a significantly larger number of LV than either H&E or LYVE-1/MCAB (P<0.0001). LYVE-1/PCAB detected LVI in 25/67 cases (37.3%) and their presence was significantly associated with both lymph node metastasis (chi(2)=4.698, P=0.0248) and unfavourable overall survival (OS) (P=0.0453), while not relapse- free survival (RFS) (P=0.2948). LMVD had no influence for RFS and OS (P=0.4879, P=0.1463, respectively). Our study demonstrates that immunohistochemistry with LYVE-1/PCAB is a highly sensitive method for detecting tumour LV/LVI in breast cancer and LVI is a useful prognostic indicator for lymphatic tumour dissemination.

Incarcerated transdiaphragmatic intercostal hernia preceded by Chilaiditi's syndrome.

Transdiaphragmatic hernia most often develops after blunt or penetrating thoracoabdominal trauma. We report on the case of a 73-year-old man who underwent emergency ileocoecal resection for an incarcerated transdiaphragmatic intercostal hernia. The patient's history included both a lumbotomy for right nephrectomy and Chilaiditi's syndrome. The literature regarding both transdiaphragmatic intercostal herniation and Chilaiditi's syndrome is reviewed in relation to the presented case.

Distribution of lymphatic vessels in normal and arthritic human synovial tissues.

OBJECTIVES: To investigate the distribution of lymphatic vessels in normal, rheumatoid arthritis (RA) and osteoarthritis (OA) synovium. METHODS: Synovial tissues from 5 normal controls, 14 patients with RA, and 16 patients with OA were studied. Lymphatic vessels were identified by immunohistochemistry using antibodies directed against the lymphatic endothelial hyaluronan receptor (LYVE-1) and recognised blood vessel endothelial markers (factor VIII, CD34, CD31). RESULTS: Lymphatic vessels were found in all zones of the normal, OA, and RA synovial membrane. Few lymphatic vessels were seen in the sublining zone in normal and OA synovium which did not show villous hypertrophy. However, in both RA synovium and OA synovium showing villous hypertrophy and a chronic inflammatory cell infiltrate, numerous lymphatic vessels were seen in all zones of the synovial membrane, including the sublining zone of the superficial subintima. CONCLUSIONS: Lymphatic vessels are present in normal and arthritic synovial tissues and are more numerous and prominent where there is oedema and an increase in inflammatory cells in the subintima, particularly in RA. This may reflect increased transport of hyaluronan and leucocyte trafficking in inflamed synovial tissues.

Transient myelopathy of the cervical posterior columns in a young man with recently diagnosed diabetes mellitus.

We present the case of a young man with recently manifesting type I diabetes mellitus who progressively developed clinical and radiological signs and symptoms of cervical posterior column myelopathy with eventually spontaneous and complete recovery as concerns both clinical symptomatology and MRI abnormalities.

LYVE-1, the lymphatic system and tumor lymphangiogenesis.

Previous research into hyaluronan (HA) has focused on the role of this abundant tissue glycosaminoglycan in promoting cell migration through interactions with its transmembrane receptor CD44 on inflammatory leukocytes and tumor cells. The recent discovery of a new HA receptor, LYVE-1 (lymphatic vessel endothelial HA receptor), expressed predominantly in lymphatic vessels, highlights another aspect of HA biology: its continuous transit through the lymphatic system and its potential involvement in lymph node homing by CD44+ leukocytes and tumor cells. The functional role of LYVE-1 in lymphatic vessels and its application as a marker to study tumor lymphangiogenesis are important areas of investigation.

Adenoviral expression of vascular endothelial growth factor-C induces lymphangiogenesis in the skin.

The growth of blood and lymphatic vasculature is mediated in part by secreted polypeptides of the vascular endothelial growth factor (VEGF) family. The prototype VEGF binds VEGF receptor (VEGFR)-1 and VEGFR-2 and is angiogenic, whereas VEGF-C, which binds to VEGFR-2 and VEGFR-3, is either angiogenic or lymphangiogenic in different assays. We used an adenoviral gene transfer approach to compare the effects of these growth factors in adult mice. Recombinant adenoviruses encoding human VEGF-C or VEGF were injected subcutaneously into C57Bl6 mice or into the ears of nude mice. Immunohistochemical analysis showed that VEGF-C upregulated VEGFR-2 and VEGFR-3 expression and VEGF upregulated VEGFR-2 expression at 4 days after injection. After 2 weeks, histochemical and immunohistochemical analysis, including staining for the lymphatic vessel endothelial hyaluronan receptor-1 (LYVE-1), the vascular endothelial marker platelet-endothelial cell adhesion molecule-1 (PECAM-1), and the proliferating cell nuclear antigen (PCNA) revealed that VEGF-C induced mainly lymphangiogenesis in contrast to VEGF, which induced only angiogenesis. These results have significant implications in the planning of gene therapy using these growth factors.

Mouse LYVE-1 is an endocytic receptor for hyaluronan in lymphatic endothelium.

The glycosaminoglycan hyaluronan is a key substrate for cell migration in tissues during inflammation, wound healing, and neoplasia. Unlike other matrix components, hyaluronan (HA) is turned over rapidly, yet most degradation occurs not locally but within distant lymph nodes, through mechanisms that are not yet understood. While it is not clear which receptors are involved in binding and uptake of hyaluronan within the lymphatics, one likely candidate is the lymphatic endothelial hyaluronan receptor LYVE-1 recently described in our laboratory (Banerji, S., Ni, J., Wang, S., Clasper, S., Su, J., Tammi, R., Jones, M., and Jackson, D.G. (1999) J. Cell Biol. 144, 789-801). Here we present evidence that LYVE-1 is involved in the uptake of hyaluronan by lymphatic endothelial cells using a new murine LYVE-1 orthologue identified from the EST data base. We show that mouse LYVE-1 both binds and internalizes hyaluronan in transfected 293T fibroblasts in vitro and demonstrate using immunoelectron microscopy that it is distributed equally among the luminal and abluminal surfaces of lymphatic vessels in vivo. In addition, we show by means of specific antisera that expression of mouse LYVE-1 remains restricted to the lymphatics in homozygous knockout mice lacking a functional gene for CD44, the closest homologue of LYVE-1 and the only other Link superfamily HA receptor known to date. Together these results suggest a role for LYVE-1 in the transport of HA from tissue to lymph and imply that further novel hyaluronan receptors must exist that can compensate for the loss of CD44 function.

Vascular endothelial growth factor-C-mediated lymphangiogenesis promotes tumour metastasis.

Metastasis is a frequent and lethal complication of cancer. Vascular endothelial growth factor-C (VEGF-C) is a recently described lymphangiogenic factor. Increased expression of VEGF-C in primary tumours correlates with dissemination of tumour cells to regional lymph nodes. However, a direct role for VEGF-C in tumour lymphangiogenesis and subsequent metastasis has yet to be demonstrated. Here we report the establishment of transgenic mice in which VEGF-C expression, driven by the rat insulin promoter (Rip), is targeted to beta-cells of the endocrine pancreas. In contrast to wild-type mice, which lack peri-insular lymphatics, RipVEGF-C transgenics develop an extensive network of lymphatics around the islets of Langerhans. These mice were crossed with Rip1Tag2 mice, which develop pancreatic beta-cell tumours that are neither lymphangiogenic nor metastatic. Double-transgenic mice formed tumours surrounded by well developed lymphatics, which frequently contained tumour cell masses of beta-cell origin. These mice frequently developed pancreatic lymph node metastases. Our findings demonstrate that VEGF-C-induced lymphangiogenesis mediates tumour cell dissemination and the formation of lymph node metastases.

VEGF-D promotes the metastatic spread of tumor cells via the lymphatics.

Metastasis to local lymph nodes via the lymphatic vessels is a common step in the spread of solid tumors. To investigate the molecular mechanisms underlying the spread of cancer by the lymphatics, we examined the ability of vascular endothelial growth factor (VEGF)-D, a ligand for the lymphatic growth factor receptor VEGFR-3/Flt-4, to induce formation of lymphatics in a mouse tumor model. Staining with markers specific for lymphatic endothelium demonstrated that VEGF-D induced the formation of lymphatics within tumors. Moreover, expression of VEGF-D in tumor cells led to spread of the tumor to lymph nodes, whereas expression of VEGF, an angiogenic growth factor which activates VEGFR-2 but not VEGFR-3, did not. VEGF-D also promoted tumor angiogenesis and growth. Lymphatic spread induced by VEGF-D could be blocked with an antibody specific for VEGF-D. This study demonstrates that lymphatics can be established in solid tumors and implicates VEGF family members in determining the route of metastatic spread.

Induction of tumor lymphangiogenesis by VEGF-C promotes breast cancer metastasis.

Metastasis of breast cancer occurs primarily through the lymphatic system, and the extent of lymph node involvement is a key prognostic factor for the disease. Whereas the significance of angiogenesis for tumor progression has been well documented, the ability of tumor cells to induce the growth of lymphatic vessels (lymphangiogenesis) and the presence of intratumoral lymphatic vessels have been controversial. Using a novel marker for lymphatic endothelium, LYVE-1, we demonstrate here the occurrence of intratumoral lymphangiogenesis within human breast cancers after orthotopic transplantation onto nude mice. Vascular endothelial growth factor (VEGF)-C overexpression in breast cancer cells potently increased intratumoral lymphangiogenesis, resulting in significantly enhanced metastasis to regional lymph nodes and to lungs. The degree of tumor lymphangiogenesis was highly correlated with the extent of lymph node and lung metastases. These results establish the occurrence and biological significance of intratumoral lymphangiogenesis in breast cancer and identify VEGF-C as a molecular link between tumor lymphangiogenesis and metastasis.

Pachymeningitis luetica: a case report.

Pachymeningitis luetica is extremely rare in developed countries. We describe a 41-year-old male patient with pachymeningitis luetica, multiple ischaemic infarctions, and severe hydrocephalus. The delay in making the diagnosis contributed to patient's death. Rapid diagnosis is essential on the slightest suspicion of an infection by Treponema pallidum, because timely treatment with antibiotics is effective.

Early detection of sacroiliitis on magnetic resonance imaging and subsequent development of sacroiliitis on plain radiography. A prospective, longitudinal study.

OBJECTIVE: To investigate the diagnostic value of magnetic resonance imaging (MRI) in the detection of early sacroiliitis. METHODS: Twenty-five consecutive HLA-B27 positive patients with inflammatory low back pain and < or = grade 2 unilateral sacroiliitis on conventional radiography (modified New York criteria) were studied. Erythrocyte sedimentation rate, C-reactive protein, plain radiography (PR), and MRI of the sacroiliac (SI) joints were obtained at study entry and PR of the SI joints after 3 years. Each radiograph and MR image set was interpreted independently. SI joints were scored according to the modified New York Criteria for radiological sacroiliitis. MRI scans were also scored for the presence of subchondral marrow edema. The relationship between > or = grade 2 sacroiliitis (by modified New York criteria for radiological sacroiliitis) shown on MRI and the subsequent development of > or = grade 2 sacroiliitis on PR after 3 years was investigated. RESULTS: At study entry > or = grade 2 sacroiliitis was found on MRI in 36 of 50 SI joints. Edema was found in 20 of 50 SI joints. After 3 years > or = grade 2 sacroiliitis was found on PR in 21 of 44 SI joints. The positive predictive value of > or = grade 2 sacroiliitis on MRI for the development of > or = grade 2 sacroiliitis on PR after 3 years was 60%; sensitivity was 85% and specificity 47%. CONCLUSION: Our data suggest that MRI of the SI joints can be used to identify sacroiliitis earlier than PR.

Meningioma presenting as Tolosa-Hunt syndrome.

A 23-year-old woman was admitted with headache, nausea, vomiting and blurred vision on the left side. Neurological examination showed ptosis with a complete internal and external ophthalmoplegia and a red fullness around the left orbita. Computed tomographic scanning of the brain revealed no abnormalities. As she improved on high doses of steroids a diagnosis of Tolosa-Hunt syndrome (THS) seemed to be indicated. However, magnetic resonance imaging (MRI) showed a lesion with intermediate signal intensity in the left cavernous sinus. Craniotomy was performed when symptoms of THS recurred. Histopathological examination revealed a meningioma with a papillary aspect and some mitoses. This case illustrates that: (1) THS is still a diagnosis by exclusion; (2) MRI and histopathological examination are important if there is any doubt about the diagnosis; and (3) also when there is no doubt, improvement after steroid therapy may be a diagnostic pitfall. Therefore, not only MRI but also orbital phlebography and angiography should seriously be considered.

Heparan sulfate-modified CD44 promotes hepatocyte growth factor/scatter factor-induced signal transduction through the receptor tyrosine kinase c-Met.

CD44 has been implicated in tumor progression and metastasis, but the mechanism(s) involved is as yet poorly understood. Recent studies have shown that CD44 isoforms containing the alternatively spliced exon v3 carry heparan sulfate side chains and are able to bind heparin-binding growth factors. In the present study, we have explored the possibility of a physical and functional interaction between CD44 and hepatocyte growth factor/scatter factor (HGF/SF), the ligand of the receptor tyrosine kinase c-Met. The HGF/SF-c-Met pathway mediates cell growth and motility and has been implicated in tumor invasion and metastasis. We demonstrate that a CD44v3 splice variant efficiently binds HGF/SF via its heparan sulfate side chain. To address the functional relevance of this interaction, Namalwa Burkitt's lymphoma cells were stably co-transfected with c-Met and either CD44v3 or the isoform CD44s, which lacks heparan sulfate. We show that, as compared with CD44s, CD44v3 promotes: (i) HGF/SF-induced phosphorylation of c-Met, (ii) phosphorylation of several downstream proteins, and (iii) activation of the MAP kinases ERK1 and -2. By heparitinase treatment and the use of a mutant HGF/SF with greatly decreased affinity for heparan sulfate, we show that the enhancement of c-Met signal transduction induced by CD44v3 was critically dependent on heparan sulfate moieties. Our results identify heparan sulfate-modified CD44 (CD44-HS) as a functional co-receptor for HGF/SF which promotes signaling through the receptor tyrosine kinase c-Met, presumably by concentrating and presenting HGF/SF. As both CD44-HS and c-Met are overexpressed on several types of tumors, we propose that the observed functional collaboration might be instrumental in promoting tumor growth and metastasis.

Early onset cerebellar ataxia with retained tendon reflexes: foot deformity in a first grade family member.

Early onset cerebellar ataxia with retained tendon reflexes (EOCA) is a clinical syndrome characterised by progressive cerebellar ataxia with an onset before the age of 25 years and a wide spectrum of associated features. It is distinguished from Friedreich's ataxia (FA) mainly by the preservation of tendon reflexes, a better prognosis, and the absence of GAA expansion in the frataxin gene. Although EOCA is thought to be a hereditary disorder with an autosomal recessive mode of inheritance, genetic heterogeneity might underlie the spectrum of clinical features. In this case report we describe a patient with EOCA accompanied by pes cavus, hammer toes and peripheral neuropathy. The patient's father did not have any ataxia, but had the same foot deformities as his daughter and a slight peripheral neuropathy. The possible relationship between these clinical features is discussed.

Extended extradural spinal arachnoid cyst: an unusual cause of progressive spastic paraparesis.

A 15-year old girl presented with a slowly progressive spastic paraparesis since the age of 12. Creatine kinase was slightly increased. Muscle biopsy carried out during tendon surgery for severe toe-walking showed 'myopathic' changes. Subsequent neurological evaluation and radiological studies revealed a large extradural arachnoid cyst extending from the 11th thoracic vertebra to the first lumbar vertebra. Her condition improved after operation. The 'myopathic' features turned out to be the result of chronic spinal compression. MRI is the method of choice to examine patients with non-hereditary progressive spastic paraparesis. Muscle biopsy and tendon surgery should not be performed, without careful neurological examination.

MRI locates a posterior fossa aneurysm in the fourth ventricle.

We report a rare case of a ruptured aneurysm of the choroidal branch of the left posterior inferior cerebellar artery (PICA) located in the fourth ventricle. Digital subtraction angiography revealed this PICA aneurysm but the exact location remained unknown. The unique location in the fourth ventricle was subsequently shown by magnetic resonance imaging (MRI). The patient died and the final diagnosis was confirmed by autopsy. To our knowledge, this is one of the few reported cases of a PICA aneurysm in the fourth ventricle and the only one which was confirmed by the combination of MRI and autopsy.

Primary cerebellar nocardiosis and alveolar proteinosis.

We report a patient with known asymptomatic pulmonary alveolar proteinosis (PAP) who developed a cerebellar gait disorder and dysarthria caused by an isolated cerebellar nocardial abscess. To our knowledge only 1 patient with PAP and isolated central nervous system nocardia infection has previously been reported. In this early report, diagnosis was established at autopsy. In our patient the clinical and MRI examinations of this cerebellar abscess are described and specific features leading to earlier diagnosis and successful treatment are presented.

Spondylodiscitis in ankylosing spondylitis, inflammation or trauma? A description of six cases.

An uncommon, but well recognised complication of Ankylosing Spondylitis (AS) is spondylodiscitis, a destructive discovertebral lesion also called the Andersson lesion. We describe six cases, with variable clinical presentation and radiological appearance. Two had multiple lesions, in one patient spondylodiscitis was the presenting symptom of AS. None had a history of even a minor trauma. Prognosis was good with conservative treatment including NSAID's, rest, and physiotherapy. In two cases histopathology was studied and suggested sterile inflammation as the main etiologic factor. The literature is reviewed regarding the mechanisms that may contribute to these lesions: mainly inflammatory like increasing enthesopathy or mainly mechanical like pseudoarthrosis about a fracture site. It may be that both mechanisms can result in similar destructive intervertebral disc lesions.