[Participation and support of clinical studies and other scientific investigations. Statement of the German Society for Pathology]. / Beteiligung und Unterstützung klinischer Studien und anderer wissenschaftlicher Untersuchungen. Stellungnahme der Deutschen Gesellschaft für Pathologie e. V.

Clinical studies and preclinical investigations are essential in order to test new therapies and diagnostics with the aim of sustained improvement in the treatment of patients. Fortunately, the number of clinical studies is continuously increasing and pathology and tissue-based research are included more often. The German Society for Pathology (DGP) and the pathologists it represents want to and can support this process and our clinical partners as best as possible as an equal partner. With our technologies and our specific expertise we can make a substantial contribution to the quality and the success of preclinical investigations, clinical studies and implementation of the results into clinical pathological diagnostics. In order to support this process the DGP has formulated a statement on the participation and support of clinical studies and other scientific investigations.

The antidepressant rolipram suppresses cytokine production and prevents autoimmune encephalomyelitis.

In multiple sclerosis (MS) and its animal model experimental autoimmune encephalomyelitis (EAE) the cytokines tumour necrosis factor-alpha (TNF), lymphotoxin-alpha (LT), and interferon-gamma (IFN-gamma) are of central pathogenetic importance. A therapy capable of stopping neurological deterioration in MS patients is not yet available. Here, we report that rolipram, a selective type IV phosphodiesterase inhibitor, stereospecifically suppresses the production of TNF/LT and less strongly also IFN-gamma in human and rat auto-reactive T cells. Moreover, we show that rolipram is an effective treatment for EAE. Rolipram has extensively been studied in humans for the treatment of depression, but has not yet been marketed. The data presented here identify rolipram as potential therapy for multiple sclerosis and provoke the immediate initiation of clinical trials.

Erythropoietin receptor expression in normal and neoplastic choroid plexus.

BACKGROUND: The erythropoietin receptor (EpoR) is expressed widely throughout the human CNS, including the choroid plexus. Recent studies have shown that EpoR is also expressed in various human tumors, including carcinomas, meningiomas and gliomas. Thereby, the Epo-EpoR pathway plays a role in inhibition of apoptosis and tumor growth, infiltration, angiogenesis and metastasis as well as treatment resistance and is a potential target in oncological treatment. Lower levels of EpoR have been associated with shorter survival in high grade gliomas and higher risk of tumor recurrence in meningiomas. METHODS: Since the EpoR status in human choroid plexus tumors (CPT) is not known, we investigated 57 CPT from 43 cases including 14 recurrent tumors and compared them with 23 samples of normal choroid plexus (CP). CPT samples consisted of choroid plexus papillomas/CPP (n = 41), atypical CPP (n = 15) and choroid plexus carcinoma/CPC (n = 1). EpoR expression was determined by immunohistochemistry using semi-quantitative scoring for staining intensity and was validated in exemplary cases using western blot and RT-PCR. RESULTS: EpoR expression was observed in all samples of normal and neoplastic CP with significantly lower expression levels in CPT (p < 0.001). CONCLUSION: No significant correlation was found between EpoR expression and age, gender, WHO grade, number of mitosis or tumor recurrence. EpoR expression in CPT is in line with its expression in normal CP and with previous reports on EpoR expression in other glial neoplasms. Association of EpoR levels in CPT with survival, as known in astrocytic gliomas, remains to be determined.

Secreted protein, acidic and rich in cysteine (SPARC) expression in astrocytic tumour cells negatively correlates with proliferation, while vascular SPARC expression is associated with patient survival.

AIMS: Secreted protein, acidic and rich in cysteine (SPARC) is a regulator of cell-matrix interaction and has been associated with tumour stage and patient survival in various malignancies. As no large-scale study has yet been undertaken, we investigated human brain and astrocytomas for SPARC expression and associations with tumour grade, proliferation, vascular density and patient survival. METHODS: A spectrum of 188 WHO grade I-IV astrocytic tumours and 24 autopsy cases were studied by immunohistochemistry for SPARC, MIB-1 proliferation index and CD31-positive vessels. SPARC protein expression was confirmed by quantitative real-time polymerase chain reaction and Western blot in 13 cases. RESULTS: In normal brain, SPARC is expressed in cortical marginal glia, cerebellar Bergmann glia and focally in white matter but is absent in neurones or vessels. High SPARC expression levels in the cytoplasm of astrocytic tumour cells decreased with the grade of malignancy but showed an increase with grade of malignancy in tumour vessels. SPARC negatively correlated with tumour proliferation but not with vascular density. While cytoplasmic SPARC staining was not associated with survival, vascular SPARC showed a significant association in the group of grade II-IV tumours (P = 0.02) and also in grade II astrocytomas alone (P = 0.01) with vascular SPARC associated with worse prognosis. CONCLUSIONS: SPARC is highly expressed in astrocytomas and decreases with tumour progression. We confirm an association of increased SPARC expression and decreased proliferation. While there is no association between the level of SPARC in the tumour cells and patient survival, increased tumour vascular SPARC expression is associated with decreased patient survival.

Spatio-temporal deleted in colorectal cancer (DCC) and netrin-1 expression in human foetal brain development.

AIMS: Deleted in colorectal cancer (DCC) and its ligand netrin-1 are known as axonal guidance factors, being involved in angiogenesis, migration and survival of precursor cells in the embryonic mammalian central nervous system (CNS). So far, little is known about the distribution of those molecules in human CNS development. METHODS: We investigated 22 human foetal brain specimens (12th and 28th week of gestation) for DCC and netrin-1 expression by means of immunohistochemistry, immunofluorescence and confocal laser microscopy. Statistical analysis was performed by applying a semi-quantitative score, including staining intensity and frequency and correlation with foetal age. RESULTS: DCC and netrin-1 were differentially expressed throughout the developing human foetal telencephalic and cerebellar cortical layers. Netrin-1 exhibited the highest levels in telencephalic germinal layers, whereas the strongest DCC immunoreactivity was seen in the developing cortical plate. Netrin-1 and DCC were predominantly present on cerebellar external granule layer cells. Distinct co-expression was seen in maturing foetal brainstem nuclei, cerebellar external granular layer and the choroid plexus. In contrast, endothelial cells showed strong netrin-1 expression with subsidiary DCC immunoreactivity. Pontine and telencephalic axonal fibre tracts also demonstrated strong netrin-1 expression. CONCLUSIONS: We show that DCC and netrin-1 are ubiquitously expressed in the human foetal brain; however, both exhibit a distinct spatio-temporal expression pattern. Together with the data from animal experiments, our findings might indicate also an important role for DCC and netrin-1 in human foetal CNS development.

Cerebral low-grade lymphoma and light chain deposition disease: exceedingly high IgG levels in the cerebrospinal fluid as a diagnostic clue.

Herein, we report the case of a 72-year-old male with an exceedingly rare manifestation of a low-grade lymphoma in the brain associated with light chain deposition disease (LCDD). The patient presented with epileptic seizures. Magnetic resonance imaging (MRI) of the brain revealed multiple hyperintense lesions in the right parietal lobe that were suspicious of vasculitis, low-grade glioma, or neurosarcoidosis. In the cerebrospinal fluid (CSF), but not in the serum, highly elevated IgG was found. A stereotactic biopsy of one cerebral lesion was performed. Histopathology revealed a low grade lymphoplasmacytic B-cell lymphoma with light chain deposition disease (LCDD). Bone marrow biopsy and laboratory workup did not show any systemic involvement. LCDD exclusively affecting the brain is an exceedingly rare finding. It can be associated with low-grade B-cell lymphoma. This is the first report of LCDD exclusively affecting the brain in an elderly patient. Compared with the two younger patients previously reported, the course of the disease was of a slow-evolving nature. In constellations of highly elevated IgG in CSF and multiple white matter lesions, LCDD should be considered as underlying pathology.

Diagnostic value of WT1 in neuroepithelial tumours.

AIMS: Currently, clinical trials using WT1 (Wilms tumour gene) peptide vaccines are conducted in haematopoietic malignancies and solid cancers. Single reports showed that the Wilms tumour gene product WT1 is also expressed in astrocytic neoplasms. Our aim was to investigate WT1 expression in a large cohort of various neuroepithelial tumours of different World Health Organization (WHO) grades and in normal central nervous system (CNS) tissue specimens to test its potential value as a diagnostic marker. METHODS: Specimens were assessed by RT-PCR, Western blotting and immunohistochemistry. The samples investigated in our study consisted of 334 human neuroepithelial tumours, among those 33 oligodendrogliomas, 219 astrocytomas (including 105 glioblastomas) and 47 ependymomas. RESULTS: Our results showed a de novo WT1 expression in neuroepithelial tumours. In diffuse astrocytomas and ependymomas, WT1 expression increased significantly with the grade of malignancy. In contrast, no significant difference was seen between WHO grade-II and -III oligodendrogliomas. Controlling for WHO grade, the comparison of oligodendrogliomas with ependymal and astrocytic tumours showed higher expression values for the latter. CONCLUSIONS: Our study shows that WT1 is expressed de novo in numerous neuroepithelial tumours and increases with the grade of malignancy. These results suggest an important role of WT1 in tumourigenesis and progression in human brain tumours.

Erythropoietin receptor is expressed in meningiomas and lower levels are associated with tumour recurrence.

AIMS: The Epo-EpoR pathway plays a role in tumour growth, metastasis and treatment resistance and is a potential target in oncological treatment. As the EpoR status in human meningiomas is unknown, our aim was to characterize EpoR expression in these tumours. METHODS: We examined 131 meningioma samples of all WHO grades from 116 patients by immunohistochemistry for EpoR. Among these, 25 meningiomas showed brain invasion and 29 patients had a further tumour recurrence. A group of 20 patients without tumour recurrence served as controls. In 12 cases we were able to compare both the primary and the following recurrent tumours. The presence of EpoR in meningiomas was confirmed by RT-PCR and Western blot. RESULTS: EpoR was expressed in all meningiomas. Statistical analysis revealed that the mean expression levels of EpoR were significantly lower in primary tumours with known recurrence compared with a recurrence-free control group. Additional matched pair analysis in individual cases showed no significant differences between primary and recurrent tumours. No significant correlation between EpoR expression and WHO grade, age, sex or brain invasion was detected. Using specific primer pairs for RT-PCR, we were able to detect all three known isoforms of EpoR: the full-length isoform EpoR-F, the truncated isoform EpoR-T and the soluble isoform EpoR-S. CONCLUSIONS: Our results demonstrate the expression of EpoR in meningiomas. Lower EpoR mean levels might be a useful marker for a higher recurrence risk, but further studies are needed to clarify the influence of EpoR on recurrences and the role of the different isoforms.

Benign meningioma developing late lung metastases: case report and review of the literature.

Here we report the case of a 65-year-old female with a histologically benign parietal falcine meningioma who developed multiple lung metastases 15 years after tumor resection. The meningioma was initially incompletely resected due to invasion of the sagittal sinus. Since it was diagnosed as a benign meningothelial meningioma Grade I WHO, the residual tumor was followed with serial imaging without adjuvant treatment. The patient subsequently developed lung lesions later identified as metastases. The lung lesions were successfully removed surgically and histologically diagnosed as meningothelial meningioma Grade I WHO. A repeat brain MRI revealed the known residual meningioma with no signs of interval tumor growth, but did demonstrate occlusion of the sagittal sinus. In the further course, the residual meningioma was completely removed. A review of the literature revealed only 15 well-documented cases of benign meningiomas that metastasized in an interval of up to 12 years after primary tumor resection. This case illustrates that histologically benign meningiomas Grade I WHO with stable disease of the primary tumor have the potential to develop hematogenous metastases even after a long time interval.

Concepts and developments in peripheral nerve surgery.

Nerve injuries may result in sensory and motor deficits when not treated appropriately. Especially the surgical management of nerve defects still represents a challenge for the surgeon. In these cases the grafting of autologous nerves represents the only reasonable approach. Due to the side effects associated with this method (sacrifice of donor nerves, neuroma formation in the harvesting area, limited availability of donor nerves, etc.), numerous alternatives were proposed in order to avoid the transplantation of autologous tissue. This review provides a general view on the state of the art of how to supply gaping injuries in the peripheral nerve. Furthermore new approaches emphasizing tubulization techniques for the reconstruction of lost nerve tissue are described with a special focus on various materials with their advantages and disadvantages.

Lesional accumulation of RhoA(+) cells in brains of experimental autoimmune encephalomyelitis and multiple sclerosis.

AIM: Infiltration of autoantigen-specific T cells and monocytes into the central nervous system is essential for the development of both experimental autoimmune encephalomyelitis (EAE) and multiple sclerosis (MS). RhoA is one of the best-known members of Rho GTPases, and inhibition of RhoA has been shown to attenuate the progression of EAE. The aim of this study was to investigate the expression of RhoA in brains of EAE rats and MS tissue. METHODS: EAE was induced by immunization with the synthetic peptide gpMBP68-84 in rats, and clinical severity was scored. RhoA expression pattern was investigated in brains of EAE rats at different time points and in different lesions of brain tissue specimens from six MS brains and five neuropathologically unaffected controls by immunohistochemistry. METHODS: In EAE rat brains, accumulation of RhoA(+) cells reached maximal levels around Day 13, correlating to the clinical severity of EAE, and up-regulation lasted until the recovery stage of the disease. Double-labelling experiments showed that the major cellular sources of RhoA were reactive macrophages/microglia. While RhoA(+) cells in normal human brain parenchyma were rarely observed, RhoA expression was found to be spatially associated with MS lesions, showing a marked decrease from active lesions via chronic stages to its near absence in normal-appearing white matter. In addition, major RhoA(+) cells in brain parenchyma of MS were identified to be activated macrophages/microglia. CONCLUSION: Our present data indicated that RhoA may play an important role during the effector phase of EAE and MS. Therefore, RhoA inhibitors might be a therapeutic option for MS patients.

The clinico-surgico-pathological spectrum of myxopapillary ependymomas--report of four unusal cases and review of the literature.

According to the WHO grading system, myxopapillary ependymomas are assigned to WHO Grade I. However, the clinico-pathological spectrum might be very heterogenous. Herein, we report 4 cases exhibiting lumbar tumor masses, 1 causing muscular atrophy over a 30-year period, 3 displaying clinical history of persisting lumbar pain for only several weeks. All tumors were crooked with dura and spinal roots resulting in incomplete resection in three cases. On histological examination, two tumors were almost acellular and showed polycyclic hyaline and fibrotic extracellular matrix leading to differential diagnoses of chordoma, meningioma, fibrolipoma and ependymoma. Finally, together with the immunohistochemical investigations, electron microscopy led to the diagnosis of myxopapillary ependymoma, WHO Grade I, with massive degenerative changes. The other 2 cases presented with the typical neuropathology of myxopapillary ependymomas but showed local recurrence within 1 and 13 years throughout the whole neuraxis, and in 1 case additional metastases of the 3rd ventricle. Although the morphological feature of these myxopapillary ependymomas was benign, the presented cases showed that the biological behavior of myxopapillary tumors might differ greatly and that these tumors present a serious operative and diagnostic challenge. Myxopapillary ependymomas occur most often in the lumbosacral region. Due to the anatomic complexity of the cauda equina, a complete resection can be technically challenging in this region. However, a gross total resection at the primary surgery is the most predictive factor for the outcome.

Primary anaplastic ganglioglioma with a small-cell glioblastoma component.

Gangliogliomas usually present as benign tumors corresponding to World Health Organization (WHO) Grade I. Very rarely, gangliogliomas show histological features of malignancy and are then classified as anaplastic gangliogliomas of WHO Grade III or IV. In most cases, anaplastic gangliogliomas developed after radiation therapy or progression from a pre-existing low-grade ganglioglioma. Here, we report the case of a 77-year-old male patient who was operated on a primary ganglioglioma with a highly anaplastic glial component corresponding to a small-cell glioblastoma. To our knowledge, this is the first reported case of a primary anaplastic ganglioglioma with a small-cell glioblastoma component.

Atypical type II silent corticotrophic adenoma developing into Cushing's disease upon second recurrence.

Herein, we report the case of a 73-year old male patient who presented with two recurrences of a pituitary adenoma within a period of 15 years. The first tumor resection 15 years ago revealed a non-functioning pituitary macroadenoma. 11 years later, the first recurrence of the tumor was reoperated. Throughout the early course of the disease, he suffered from secondary adrenal insufficiency and required replacement therapy with hydrocortisone. Currently, he presented with the second recurrence and clinical examination revealed signs of Cushing's disease. This was clearly confirmed by endocrinological evaluation. A retrospective analysis of all histological and immunohistochemical slides rendered an adenoma exhibiting chromophobia, ACTH-positivity and features of atypia such as elevated p53 and Ki67 expression as well as nuclear polymorphism. According to the revised WHO classification it was classified as atypical type II silent corticotroph adenoma at the time of the first and second surgery. The specimen removed during the recent surgery displayed the same histological features and was classified as corticotroph adenoma. The combination of an atypical type II adenoma and the switch in the hormone status to an endocrinologically active adenoma makes this case exceedingly rare.

Adult medulloblastoma: prognostic factors and response to therapy at diagnosis and at relapse.

Adult medulloblastoma is a rare tumor with few retrospective studies published so far. The role of adjuvant chemotherapy or chemotherapy at relapse is unclear. This study reports therapy and outcome in all adult (>or=16 years old) medulloblastoma (n=34) and supratentorial primitive neuroectodermal tumor (PNET) patients (n=2) treated in 2 neuro-oncological centers between 1976 and 2002. The median age was 24.5 years (range 16-76). After resection, 16 patients were treated with craniospinal radiotherapy alone, 20 patients also received adjuvant chemotherapy (8 vincristine, CCNU, cisplatin; 7 methotrexate alone or methotrexate/vincristine-based polychemotherapy; 5 other protocols). Median survival in the whole cohort was 126 months (2+ - 200+months). Five-year and 10-year survival rates were 79 % and 56%. Adjuvant chemotherapy was associated with a non-significant trend to prolonged survival (relative risk (RR) 1.89; p=0.068). The median progression-free survival (PFS) after primary therapy was 83 months. At relapse, 10 of 12 evaluable patients achieved a complete response upon second-line therapy. The median survival times from first (n=17) and second relapse (n=9) were 21 months (0-67+ months; 5/17 without second relapse) and 20 months (1-29 months). Cox regression analysis revealed the infiltration of the floor of the 4(th) ventricle at diagnosis as the only therapy-independent prognostic factor (RR 0.48; p=0.03). In conclusion, adjuvant chemotherapy may prolong survival in adult medulloblastoma patients. Moreover, second-line therapy may be beneficial for these patients. As in pediatric medulloblastoma patients, primary infiltration of the floor of the 4(th) ventricle indicates a poor prognosis.

Allograft inflammatory factor-1 is expressed by macrophages in injured skeletal muscle and abrogates proliferation and differentiation of satellite cells.

Secretion of regulatory peptides by macrophages in injured skeletal muscle constitutes a pivotal determinator of tissue homeostasis. We analyzed expression of a novel Ca2+- binding peptide expressed by activated macrophages, the allograft inflammatory factor-1 (AIF-1), in rat devascularized skeletal muscle. AIF-1 expression was observed in 94% of all macrophages at the site of the injury 48 hours postdevascularization. The physiological function of AIF-1 in injured skeletal muscle was analyzed using a rat in-vitro model of satellite cell proliferation and differentiation. Addition of AIF-1 to the culture medium resulted in a concentration-dependent and reversible reduction of the total number of cells expressing M-cadherin (p < or = 0.0001), a mediator of the differentiation process of skeletal muscle cells, the proliferation associated PCNA (p < or = 0.0001), and the initiator of muscle differentiation myogenin (p < or = 0.0001). These results provide convincing evidence that activated AIF-1 expressing macrophages constitute the predominant cell type in skeletal muscle 48 hours postinjury, and that AIF-1 regulates reduced proliferation, differentiation, and activation of satellite cells.

In vivo expression of insulin-like growth factor-binding protein-2 in human gliomas increases with the tumor grade.

Human central nervous system tumors and glioma cell lines highly express the insulin-like growth factor-binding protein (IGFBP)-2. As IGFBP-2 can affect tumor growth, we studied the relationship between IGFBP-2 expression and the malignancy of brain tumors in vivo. To do so, we investigated by immunohistochemistry the accumulation of IGFBP-1, -2, and -3 in 50 human gliomas classified by the WHO Malignancy Scale. Double labeling using anti-CD68 (monocytes/macrophages), antiglial fibrillary acidic protein, and anti-CD3 (T cells) antibodies was performed to further characterize the IGFBP-1, -2, and -3(+) cells. The expression of IGFBP messenger RNAs (mRNAs) was tested by RT-PCR in tumor samples from nine gliomas of different grades and in eight cell lines representing the cellular composition of human glioma. As controls, the accumulation of IGFBP-2 was investigated in normal brain and in the rat C6 glioblastoma model. IGFBP-1 and -3 accumulated in endothelial and macrophage/microglial cells. IGFBP-2(+) macrophage/microglial and glioma cells clustered in the immediate vicinity of focal necrosis of the human gliomas as well as of the rat C6 glioblastoma. The labeling score of IGFBP-1 accumulation in endothelial cells correlated negatively (P: = 0.0229), and that of IGFBP-2 accumulation in glioma cells correlated positively (P: < 0.0006) with the tumor grade of the gliomas. In addition, RT-PCR analysis confirmed mRNA expression of IGFBP-1, -2, and -3 by the gliomas and glial cells. Small amounts of IGFBP-1 and -3 mRNA, but high amounts of IGFBP-2 mRNA, were detectable in macrophage-like and glioma cell lines. The results suggest cell type-specific accumulation of IGFBP-1, -2, and -3 in human glial tumors of the brain. The increase in IGFBP-2 expression with this malignancy suggests a role of IGFBP-2 in the biology of human gliomas.

Interaction of T lymphocytes with cerebral endothelial cells in vitro.

As a prerequisite of inflammatory lesion formation in (auto-)immune disease of the central nervous system, lymphocytes have to interact with brain endothelia. In recent years much progress has been made towards a better understanding of mechanisms and factors involved in organ specific homing of lymphocytes. Many lines of evidence indicate that T lymphocytes recognizing antigens which are exclusively beyond the blood-brain barrier cross this barrier only when they are in an activated state, irrespective of their antigen specificity. Antigen presentation by blood-brain barrier endothelia, however, may play a role in later stages of florid inflammation.

Bag-1 and Bcl-2 gene transfer in malignant glioma: modulation of cell cycle regulation and apoptosis.

Bag-1 is a heat shock 70 kDa (Hsp70)-binding protein that can collaborate with Bcl-2 in suppressing apoptosis under some conditions. Here, we report that 11 of 12 human glioma cell lines express Bag-1 protein in vitro. Moreover, 15 of 19 human glioblastomas expressed Bag-1 as assessed by immunohistochemistry in primary tumor specimens. To examine the biological effects of Bag-1 in glioma cells, we expressed Bag-1 or Bcl-2 transgenes in 2 human malignant glioma cell lines, LN-18 and LN-229. Bag-1 significantly slowed glioma cell growth and reduced clonogenicity of both cell lines in vitro. Coexpressed Bcl-2 abrogated these effects of Bag-1. Intracranial LN-229 glioma xenografts implanted into nude mice revealed a substantial growth advantage afforded by Bcl-2. Bag-1 had no such effect, either in the absence or presence of Bcl-2. Upon serum starvation in vitro, Bcl-2 prevented cell death whereas Bag-1 did not. Both Bcl-2 and Bag-1 slowed proliferation of serum-starved cells when expressed alone. Importantly, coexpression of Bcl-2 and Bag-1 provided a distinct growth advantage under conditions of serum starvation that is probably the result of (i) the death-preventing activity of Bcl-2 and (ii) the property of Bag-1 to overcome a Bcl-2-mediated enhancement of exit from the cell cycle. In contrast to these Bcl-2/Bag-1 interactions observed under serum starvation conditions, Bag-1 did not further enhance the strong protection from staurosporine-, CD95 (Fas/Apo1) ligand-, Apo2 ligand (TRAIL)- or chemotherapeutic drug-induced apoptosis afforded by Bcl-2. Taken together, these results indicate a role for Bag-1/Bcl-2 interactions in providing a survival advantage to cancer cells in a deprived microenvironment that may be characteristic of ischemic/hypoxic tumors such as human glioblastoma multiforme, and suggest that Bcl-2/Bag-1 interactions also modulate cell proliferation.

Expression and functional activity of heat shock proteins in human glioblastoma multiforme.

OBJECTIVE: To assess the expression of heat shock proteins (HSP) in glioma cells in vitro and in vivo and to examine their role in resistance to apoptosis. BACKGROUND: HSP are expressed in response to various forms of stress. Constitutive HSP expression may confer resistance to cytotoxic stimuli in human cancers. METHODS: HSP expression was assessed by immunoblot analysis in glioma cells in vitro and by immunocytochemistry in human glioblastomas in vivo. Modulation of apoptosis by hyperthermia-mediated HSP induction was examined in glioma cell lines in vitro. RESULTS: Immunoblot analysis revealed constitutive expression of HSP27, HSP72, HSP73, and HSP90 in all 12 human glioma cell lines. B-crystallin (alphaBC) was expressed in 3 of 12 cell lines. High levels of alphaBC and HSP72 correlated with drug resistance and high p53 levels in vitro. Transient hyperthermia (43 degrees C/2 hours) induced HSP27 and HSP72 expression but had no effect on the levels of alphaBC, HSP73, or HSP90. HSP induction provided no survival advantage against subsequent cytotoxic challenges, including cytotoxic cytokines and radiochemotherapy. Immunohistochemistry showed strong expression of all HSP in vivo. The comparative analysis of HSP27, alphaBC, HSP72, HSP73, and HSP90 expression in 24 paired samples of first resections and recurrences of human glioblastoma multiforme revealed no impact of HSP expression on response to adjuvant radiochemotherapy and no modulation of HSP expression by radiochemotherapy. CONCLUSIONS: High constitutive, as opposed to inducible, expression of HSP may play a role in the primary resistance of human malignant gliomas to cytotoxic radiochemotherapy. Superinduction of HSP levels by hyperthermia in vitro provided no further survival advantage.