Depression and quality of life in adults perceiving exposure to parental alienation behaviors.

BACKGROUND: The current study is aimed at examining the relationship between exposure to parental alienation (PA) behaviors, depression, and health-related quality of life (HRQoL) in Italian adults. METHODS: Four hundred ninety-one adults were tested. Participants filled out the following self-rating scales: The Baker Strategy Questionnaire (BSQ), the Beck Depression Inventory - II (BDI-II) and its brief version (6-item version of the BDI-II), the Short-Form 36 (SF-36) Health Survey for measuring HRQoL and its brief version including 3 items (WHO-3) of the 5-item World Health Organization Well-Being Index. RESULTS: Findings revealed statistically significant differences between participants who reported PA and those who did not. Participants who reported exposure to PA behaviors had higher scores on the original BDI-II and its 6-item version (p < 0.05, p < 0.01, respectively); they had also lower levels of HRQoL as resulting from 6 of the 8 SF-36 domains (at least p < 0.05), including lower scores on the WHO-3 (p < 0.01). Perceiving an exposure to PA behaviors significantly increased the likelihood of being above the clinical cut-off on the BDI-II (p < 0.01), the 6-item version of the BDI-II (p < 0.05), and the WHO-3 (p < 0.05). Moreover, perceiving an exposure to PA increased the odds of diminished HRQoL (OR = 2.43 and OR = 1.92 for general health and social functioning domains, respectively). CONCLUSIONS: Childhood exposure to PA was related to higher likelihood of depressive symptoms and diminished HRQoL in adulthood. Our findings suggest the need for preventive and clinical interventions to protect vulnerable children involved in PA from negative outcomes.

Identification of de novo mutations and rare variants in hypoplastic left heart syndrome.

Hypoplastic left heart syndrome (HLHS) is one of the most severe congenital heart malformations, characterized by underdevelopment of the structures in the left heart-aorta complex. The majority of cases are sporadic. Although multiple genetic loci have been tentatively implicated in HLHS, no gene or pathway seems to be specifically associated with the disease. To elucidate the genetic basis of HLHS, we analyzed 53 well-characterized patients with isolated HLHS using an integrated genomic approach that combined DNA sequencing of five candidate genes (NKX2-5, NOTCH1, HAND1, FOXC2 and FOXL1) and genome-wide screening by high-resolution array comparative genomic hybridization. In 30 patients, we identified two novel de novo mutations in NOTCH1, 23 rare patients inherited gene variants in NOTCH1, FOXC2 and FOXL1, and 33 rare patients mostly inherited copy-number variants. Some of the identified variations coexisted in the same patient. The biological significance of such rare variations is unknown, but our findings strengthen the role of NOTCH pathway in cardiac valve development, indicating that HLHS is, at least in part, a 'valve' disease. This is the first report of de novo mutations associated with isolated HLHS. Moreover, the coexistence of multiple rare variants suggests in some cases a cumulative effect, as shown for other complex disease.

Precursory worldwide signatures of earthquake occurrences on Swarm satellite data.

The study of the preparation phase of large earthquakes is essential to understand the physical processes involved, and potentially useful also to develop a future reliable short-term warning system. Here we analyse electron density and magnetic field data measured by Swarm three-satellite constellation for 4.7 years, to look for possible in-situ ionospheric precursors of large earthquakes to study the interactions between the lithosphere and the above atmosphere and ionosphere, in what is called the Lithosphere-Atmosphere-Ionosphere Coupling (LAIC). We define these anomalies statistically in the whole space-time interval of interest and use a Worldwide Statistical Correlation (WSC) analysis through a superposed epoch approach to study the possible relation with the earthquakes. We find some clear concentrations of electron density and magnetic anomalies from more than two months to some days before the earthquake occurrences. Such anomaly clustering is, in general, statistically significant with respect to homogeneous random simulations, supporting a LAIC during the preparation phase of earthquakes. By investigating different earthquake magnitude ranges, not only do we confirm the well-known Rikitake empirical law between ionospheric anomaly precursor time and earthquake magnitude, but we also give more reliability to the seismic source origin for many of the identified anomalies.

Rescue of motoneurons from cell death by a purified skeletal muscle polypeptide: effects of the ChAT development factor, CDF.

Rat skeletal muscle contains a 22 kd polypeptide that increases the level of choline acetyltransferase (ChAT) activity in cultures of embryonic rat spinal cord neurons and has been purified to homogeneity. The application of this factor, ChAT development factor or CDF, to developing chick embryos during the period of naturally occurring motoneuron cell death significantly increased the survival of motoneurons but did not affect the survival of dorsal root ganglion neurons or sympathetic preganglionic neurons (column of Terni). These results provide the first demonstration that an isolated, skeletal muscle-derived molecule can selectively enhance the survival of motoneurons in vivo and suggest that CDF may function in vivo to regulate the survival and development of motoneurons.

Heparanase expression and activity influences chondrogenic and osteogenic processes during endochondral bone formation.

Endochondral bone formation is a highly orchestrated process involving coordination among cell-cell, cell-matrix and growth factor signaling that eventually results in the production of mineralized bone from a cartilage template. Chondrogenic and osteogenic differentiation occur in sequence during this process, and the temporospatial patterning clearly requires the activities of heparin binding growth factors and their receptors. Heparanase (HPSE) plays a role in osteogenesis, but the mechanism by which it does so is incompletely understood. We used a combination of ex vivo and in vitro approaches and a well described HPSE inhibitor, PI-88 to study HPSE in endochondral bone formation. In situ hybridization and immunolocalization with HPSE antibodies revealed that HPSE is expressed in the peri-chondrium, peri-osteum, and at the chondro-osseous junction, all sites of key signaling events and tissue morphogenesis. Transcripts encoding Hpse also were observed in the pre-hypertrophic zone. Addition of PI-88 to metatarsals in organ culture reduced growth and suggested that HPSE activity aids the transition from chondrogenic to osteogenic processes in growth of long bones. To study this, we used high density cultures of ATDC5 pre-chondrogenic cells grown under conditions favoring chondrogenesis or osteogenesis. Under chondrogenic conditions, HPSE/Hpse was expressed at high levels during the mid-culture period, at the onset of terminal chondrogenesis. PI-88 addition reduced chondrogenesis and accelerated osteogenesis, including a dramatic up-regulation of osteocalcin levels. In normal growth medium, addition of PI-88 reduced migration of ATDC-5 cells, suggesting that HPSE facilitates cartilage replacement by bone at the chondro-osseous junction by removing the HS component of proteoglycans, such as perlecan/HSPG2, that otherwise prevent osteogenic cells from remodeling hypertrophic cartilage.

Neurotrophin receptors and heparanase: a functional axis in human medulloblastoma invasion.

Medulloblastoma (MB) is the most common malignant brain tumor of childhood. Although modern therapy has produced five-year survival rates as high as 70% for some MB patients, this resulted in significant long-term treatment-related morbidity. The cellular mechanisms involved in metastatic spread of medulloblastoma are largely unknown. Neurotrophins (NT) comprise a family of structurally and functionally related neurotrophic factors that are critical for central nervous system (CNS) development with nerve growth factor (NGF) being the prototypic NT. NT acts through two groups of structurally unrelated neurotrophin receptors (NTR): a family of receptor tyrosine kinases (Trks, mainly TrkA, TrkB, and TrkC) and a tumor necrosis factor receptor (TNFR)-like molecule called p75NTR TrkC expression is a good prognostic indicator for MB. TrkC binds only to neurotrophin-3 (NT-3) whereas p75 binds to all NT family members. Importantly, little is known about the biological functions of p75 in primitive neuroectodermal tumors such as MB. In contrast, NT-regulated heparanase (HPSE) is a unique extracellular matrix-degrading enzyme known to be associated with tumor progression in a wide variety of cancers. However, HPSE roles in MB invasive pathways have not been investigated. We provide evidence of a differential expression of HPSE in newly-developed medulloblastoma cell lines. Secondly, we show a correlation between HPSE expression and the invasive properties of these medulloblastoma lines. Thirdly, by performing investigations to elucidate prognostic implications of HPSE and TrkC/p75NTR expression in MB, we demonstrate a correlation between p75NTR and HPSE expression. Finally, by using antibodies specific to TrkC and immunohistochemistry (IHC) we prove that IHC scores reveal a significant expression of HPSE in 76% of MB tissues from children aged 3 years and older. Taken together, our data provide evidence that HPSE functionality, in a context linked to TrkC and p75NTR activation, may play critical roles in medulloblastoma tumor invasion and progression.

Mediation of NGF-stimulated extracellular matrix invasion by the human melanoma low-affinity p75 neurotrophin receptor: melanoma p75 functions independently of trkA.

Although overexpression of the low-affinity p75 neurotrophin receptor (p75NTR) is frequently associated with advanced stages of human melanoma progression, the functional significance of this finding is unknown. We examined whether the degree of cell surface expression of p75NTR in human melanoma cell variants determines their extent of invasion stimulated by nerve growth factor (NGF). Treatment of MeWo melanoma cells or a metastatic spontaneous wheat germ agglutinin-resistant variant subline (70W) of MeWo cells with 2.5S NGF resulted in a dose-dependent enhancement of invasion through a reconstituted basement membrane. This effect was most pronounced with the 70W subline that exhibits brain-metastasizing potential in nude mice but was not found with a poorly metastatic MeWo variant subline (3S5). The expression of p75NTR as determined by Northern blotting and immunoprecipitation analysis of 125I-labeled cell surface proteins correlated with NGF-stimulated invasion. The MeWo melanoma sublines used in this study did not express p140proto-trkA mRNA or any p140proto-trkA variant transcripts including p70trkA as determined by Northern analysis and RT-PCR analysis. Thus, these melanoma cells would not be expected to form functional p75-p140 heterodimers or p140-p140 homodimers capable of transducing an NGF-generated signal to p140proto-trkA cytoplasmic substrates. These cells did express authentic p145trkC transcripts. However, NGF did not catalytically activate p145trkC receptors via increased tyrosine phosphorylation as would be expected if p145trkC participated in the signaling established by NGF. Furthermore, a NGF-stimulated purine-analogue-sensitive kinase activity was found to coimmunoprecipitate with p75NTR. This p75NTR-associated kinase may coordinate initial signaling events evoked by p75NTR ligand interaction. Addition of 2.5S NGF, at concentrations that should saturate cell surface p75NTR, to matrix-adherent cultures of human MeWo and 70W but not 3S5 melanoma cells suppressed the expression of 92-kDa type IV collagenase and stimulated the production of 72-kDa type IV collagenase in its fully active 68-kDa form. In the absence of p140proto-trkA, the matrix-dependent effects of NGF on metalloproteinase expression of brain-metastatic 70W melanoma cells suggest a signaling role for the low-affinity melanoma p75NTR receptor and its associated purine-analogue-sensitive kinase in signaling enhanced matrix penetration of NGF-rich stromal microenvironments such as the brain.

Astrocytes contribute to the brain-metastatic specificity of melanoma cells by producing heparanase.

Neurotrophins (NTs) modulate the brain invasion of melanoma cells and the activity of an extracellular matrix degradative enzyme, heparanase, that has been recently cloned. Heparanase degrades the heparan sulfate proteoglycans (HSPGs) and is a critical mediator of tumor metastasis and angiogenesis. Because astrocytes are among the first brain cells encountered by extravasating melanoma cells, they may play important roles in the development of brain metastases. To test this hypothesis, we used purified in vitro astrocyte cultures and found that they express heparanase transcript and functional enzyme that were up-regulated by the prototypic NT, nerve growth factor. Coincubation of astrocytes (or their conditioned medium) with brain-metastatic cells resulted in a superadditive effect on heparanase activity and up to an 8-fold increase of in vitro chemoinvasion using purified HSPGs. These observations indicate that astrocytes significantly contribute to the brain colonization of melanoma cells via heparanase-driven modalities.

Flow cytometric analysis of glucocorticoid receptor using monoclonal antibody and fluoresceinated ligand probes.

Conditions were established for single cell analysis of glucocorticoid receptor (GR) content by flow cytometry using several clones of a human leukemic cell line (CCRF-CEM). These included CEM-7A, 7R, C1, and ICR 27 Tk.3 cells which were examined both by standard [3H]dexamethasone radiometric binding and by two independent flow cytometry assays. The latter involved either mouse monoclonal antibody against GR (GR-MoAb) or fluoresceinated cortisol ligand probes. For CEM-7A, 7R, and C1 cells, there was a correlation between GR-MoAb and radiometrically defined GR values. However, clone ICR-27 Tk.3 with low [3H]dexamethasone binding exhibited the highest GR-MoAb fluorescence. The fluoresceinated cortisol assay correlated with dexamethasone binding values in all four clones. Thus, GR-MoAb identifies the total immunologically reactive GR present, while the fluoresceinated cortisol assay quantifies only the functionally intact GR in terms of its initial binding. Their combined use may reveal the cellular heterogeneity of GR expression and function also in human tumor samples, to which they have been successfully applied. When coupled with DNA counterstaining, GR expression can be related directly to frequently DNA-aneuploid tumor cells and cell cycle distribution.

Neurotrophin stimulation of human melanoma cell invasion: selected enhancement of heparanase activity and heparanase degradation of specific heparan sulfate subpopulations.

Heparanase is an endo-beta-D-glucuronidase, the enzymatic targets of which are the glycosaminoglycan chains of heparan sulfate proteoglycans. Elevated levels of heparanase are associated with the metastatic potential of melanoma cells. Treatment of murine and human melanoma cells with the prototypic neurotrophin nerve growth factor (NGF) increases the production of heparanase by melanoma cells. We reported previously that physiological concentrations of NGF increased in vitro Matrigel invasion of early-passage human brain-metastatic 70W melanoma cells but not melanoma cells metastatic to other sites or nonmetastatic melanoma cells. Here we found that treatment of 70W melanoma cells with neurotrophin NT-3 increased Matrigel invasion, whereas treatment with neurotrophins other than NGF or NT-3 did not influence invasion. Mutants of NGF that do not bind to the neurotrophin receptor p75NTR or other nonneuronal growth factors were not able to enhance the invasion of 70W melanoma cells. When 70W cells were exposed to antisense oligonucleotides directed against p75NTR mRNA, there was a reduction in NGF and NT-3 binding, and the neurotrophins failed to enhance Matrigel invasion. To study the properties of heparanase in NT-regulated malignant melanoma invasive processes, we developed a sensitive heparanase assay consisting of purified [35S]heparan sulfate subpopulations separated by agarose gel electrophoresis. Incubation of 70W cells with NGF or NT-3, but not brain-derived NT factor, NT-4/5, or mutant NGF, resulted in increased release of heparanase activity that was capable of degrading a subpopulation of heparan sulfate molecules.

Abnormal expression of perlecan proteoglycan in metastatic melanomas.

Abnormal expression of proteoglycans has been implicated in cancer and metastasis primarily because these macromolecules are involved in the control of cell growth and matrix assembly. In this report, we have investigated the expression and immunolocalization of perlecan, a major heparan sulfate proteoglycan of basement membranes and pericellular matrices, in human metastatic melanomas. Twenty-six of the 27 tumor samples showed a significant increase (up to 15-fold) in the perlecan mRNA levels when compared with normal tissue. This change correlated with a vast deposition of perlecan protein core in the pericellular matrix of metastatic melanomas. Furthermore, we have established a relationship between perlecan expression in clonal melanoma cells (70W) stimulated with neurotrophins and their increased invasiveness. Interestingly, perlecan mRNA levels were up-regulated within 10 min of neurotrophin stimulation, indicating that perlecan is an early response gene. This upregulation also occurred prior to heparanase production, suggesting that perlecan expression and its regulation might play a pivotal role in the initial onset of invasion.

Stimulation of the protein tyrosine kinase c-Yes but not c-Src by neurotrophins in human brain-metastatic melanoma cells.

The c-Yes proto-oncogene (pp62c-Yes) encodes a non-receptor-type protein tyrosine kinase (NRPTK) of the Src family. c-Yes activities and protein levels are elevated in human melanoma and melanocyte cell lines. Because the neurotrophins (NT) are important in the progression of melanoma to the brain-metastatic phenotype, we determined whether NT stimulate c-Yes activity in human MeWo melanoma cells and two variant sublines with opposite metastatic capabilities, 3 S 5 and 70W. The highly brain-metastatic 70W subline had an intrinsically higher c-Yes activity than parental MeWo or poorly metastatic 3 S 5 cells. c-Yes kinase was further induced by the prototypic human NT, nerve growth factor (NGF) in a dose and time-dependent manner. In contrast, c-Src activity (pp60-Src) was similar in all these cells and unaffected by NGF exposure. Additionally, human NGF and neurotrophin-3 stimulated c-Yes in brain-metastatic 70W cells. The magnitude of c-Yes activation correlated with the degree of invasion of 70 W cells following incubation of these neurotrophins. To further examine NT stimulation of c-Yes in melanoma cells, three additional cell lines were examined. Metastatic TXM-13 and TXM-18 increased c-Yes activity in response to NGF. In contrast, no increase was observed in low-metastatic TXM-40 cells. Together, these data suggest that altered c-Yes expression may play a role in the malignant progression of the human melanocyte towards the brain-metastatic phenotype and that NT enhance the activity of c-Yes in signaling penetration into the matrix of NT-rich stromal microenvironments such as the brain.

The impact of aggressive debulking surgery and cisplatin-based chemotherapy on progression-free survival in stage III and IV ovarian carcinoma.

Forty consecutive patients with stage III and IV invasive ovarian carcinoma were treated on a phase II protocol consisting of optimal debulking surgery, induction cisplatin, cisplatin, doxorubicin, and cyclophosphamide (PAC) chemotherapy, 6-month interval laparoscopy, reinduction cisplatin, PAC chemotherapy, and second-look procedure. All 40 patients have either disease progression or have completed the 12-month protocol. Eighty-seven percent of the patients (35) underwent optimal (less than or equal to 2 cm residual) debulking surgery before chemotherapy, in spite of the fact that 50% (20) were referred to Roswell Park Memorial Institute (RPMI) as inoperable after initial surgery elsewhere. There were no postoperative deaths and chemotherapy was started in less than or equal to 14 days in 97% of the patients. Of the 40 patients, 30% (12) achieved a pathologic complete remission (11) or a clinical complete remission (one patient refused second-look surgery). The estimated 3-year survival rate was 62%, but the 3-year progression-free survival rate was only 29%. The median survival time was 48 months. The estimated 3-year progression-free survival rate was 31% for residual disease less than or equal to 2 cm. For the five patients with residual disease greater than 2 cm, four died within 3 years. The median survival time of patients with less than or equal to 2 cm residual disease was 48 months, as compared with 21 months for those with greater than 2 cm residual disease. Although the estimated 3-year survival rate of 62% is noteworthy, the 3-year progression-free survival rate of only 29% is probably indicative that in spite of extensive debulking surgery and cisplatin-based chemotherapy as used in this protocol, the long range proportion of patients "cured" will remain small.

Surgically documented response to intraperitoneal cisplatin, cytarabine, and bleomycin after intravenous cisplatin-based chemotherapy in advanced ovarian adenocarcinoma.

Thirty-one evaluable patients with stages III and IV invasive ovarian adenocarcinoma were treated on a phase II protocol of second-line intraperitoneal cisplatin, cytarabine, and bleomycin. All 31 patients received first-line intravenous (IV) cisplatin-based chemotherapy; the size of the residual cancer was documented surgically before intraperitoneal chemotherapy in all patients. Response to intraperitoneal chemotherapy was documented by a third-look laparotomy in all patients not evidencing progression of disease clinically. There were eight responses (26%): five surgical complete responses and three surgical partial responses. Responders were patients with stage III ovarian cancer, small residual disease of less than or equal to 1 cm (primarily less than or equal to 5 mm), and patients who previously had responded to cisplatin-based IV chemotherapy. Of the 15 patients with stage III ovarian cancer, residual disease less than or equal to 1 cm, and those who had responded to first-line IV cisplatin-based chemotherapy, 53% (eight) responded to second-line intraperitoneal chemotherapy. Intraperitoneal chemotherapy as used in this phase II protocol would appear to be an effective second-line treatment in advanced ovarian cancer in this specific subset of patients.

Erythropoiesis in the X-irradiated, bled rat.

Exposure to low levels of x-irradiation (50R) followed by phlebotomy (50% blood volume) one month post-irradiation, resulted in identifiable alterations of the erythropoietic status of the rat. The red cell indices revealed a decrease in mean corpuscular volume and an increase in mean corpuscular hemoglobin concentration. The half-life of 51Cr-labeled erythrocytes was 16.7 days versus 14.4 days for the untreated controls. These subjects also demonstrated hyperplastic marrows with an approximate 60% mean increase in marrow cellularity. A reproducible mortality of 25% was seen at 17 weeks post-irradiation; the one group of animals followed for an extended period exhibited an 86% cumulative mortality at 23 weeks. The observations supported the concept that x-irradiated rats exposed to a relatively low dose of X-rays (less than 10% LD/50) maintain a latent or residual injury of the bone marrow. These animals, when subsequently challenged by phlebotomy, are placed at greater risk with respect to their ability to survive.

Oncogenic role of Merlin/NF2 in glioblastoma.

Glioblastoma is the most common and aggressive primary brain tumor in adults, with a poor prognosis because of its resistance to radiotherapy and chemotherapy. Merlin/NF2 (moesin-ezrin-radixin-like protein/neurofibromatosis type 2) is a tumor suppressor found to be mutated in most nervous system tumors; however, it is not mutated in glioblastomas. Merlin associates with several transmembrane receptors and intracellular proteins serving as an anchoring molecule. Additionally, it acts as a key component of cell motility. By selecting sub-populations of U251 glioblastoma cells, we observed that high expression of phosphorylated Merlin at serine 518 (S518-Merlin), NOTCH1 and epidermal growth factor receptor (EGFR) correlated with increased cell proliferation and tumorigenesis. These cells were defective in cell-contact inhibition with changes in Merlin phosphorylation directly affecting NOTCH1 and EGFR expression, as well as downstream targets HES1 (hairy and enhancer of split-1) and CCND1 (cyclin D1). Of note, we identified a function for S518-Merlin, which is distinct from what has been reported when the expression of Merlin is diminished in relation to EGFR and NOTCH1 expression, providing first-time evidence that demonstrates that the phosphorylation of S518-Merlin in glioblastoma promotes oncogenic properties that are not only the result of inactivation of the tumor suppressor role of Merlin but also an independent process implicating a Merlin-driven regulation of NOTCH1 and EGFR.

Role of neurotrophins and neurotrophins receptors in the in vitro invasion and heparanase production of human prostate cancer cells.

The role of the neurotrophins (NTs) and their corresponding receptors (NTRs) TrkA, TrkB, TrkC, and p75NTR in neoplasia has received relatively little attention. However, because malignant cell migration within the prostate occurs predominantly by direct extension around prostatic nerves, the presence and possible upregulation of NTs from autocrine/paracrine sources and NTR expression within prostate epithelial tumor cells may be important in metastasis. We have been addressing their expression and interactions in human prostate cancer cell lines (LNCaP, PC-3, and DU145) and their role in prostate cancer invasion. In this study, we demonstrated that nerve growth factor (NGF), the prototypic NT, and NT-4/5 increased in vitro invasion through a reconstituted basement membrane and induced time- and dose-dependent expression of heparanase, a heparan sulfate-specific endo-beta-D-glucuronidase, an important molecular determinant of tumor metastasis. The NT effects were most marked in the DU 145 brain-metastatic cells and were detected at NT concentrations sufficient to fully saturate both low- and high-affinity NTRs. Additionally, we characterized the molecular expression of NT high-affinity (Trk) and low-affinity (p75NTR) receptors in these cell lines by reverse transcription-polymerase chain reaction. These lines had negligible trkA and trkC expression, although trkB was expressed in the three prostatic tumor cell lines examined. The brain-metastatic DU 145 cells were also positive for p75NTR. Our data showed that the NTs and NTRs are important in metastasis and that their expression coincides with transformation to a malignant phenotype capable of invasion along the perineural space and extracapsular metastasis to distant sites. These findings set the stage for more research into this area as related to prostate cancer evolution and may improve therapy for prostate cancer metastasis.

Inverse expression of neurotrophins and neurotrophin receptors at the invasion front of human-melanoma brain metastases.

Neurotrophins (NT), such as nerve growth factor (NGF), stimulate the growth and differentiation of several neuronal subpopulations in a distinct yet overlapping manner. Brain-metastatic human melanoma cells overexpress p75(NTR), the low-affinity neurotrophin receptor, and treatment of brain-metastatic cells with NGF stimulates extracellular matrix invasion and production of degradative enzymes in relation to the cellular expression of p75(NTR) Although human melanoma cells express high affinity neurotrophin receptors, such as TrkC (the putative receptor for NT-3), they do not express TrkA, the high-affinity NGF receptor. Using digoxigenin-labeled sense/antisense riboprobes against human p75(NTR) and NGF for in situ hybridization, we determined whether the expression of p75(NTR) and NGF mRNAs are related to brain metastasis of human melanoma. We detected p75(NTR) mRNA at the invasion front of human melanoma brain metastases, whereas p75(NTR) expression was not found in adjacent tissues. In contrast, human NGF mRNA levels were increased in tissues surrounding the melanoma lesions, supporting the notion that NGF and NT are important in determining melanoma brain-metastatic microenvironment. Using antibodies specific to p75(NTR), TrkC, NGF and related NT we found high but heterogeneous levels of p75(NTR) and TrkC expression in malignant melanomas metastatic to the brain. Lower levels of expression were found in primary melanomas or in metastatic melanomas to sites other than brain. Additionally, we found elevated levels of synthesis of NGF and NT-3 but not brain-derived neurotrophic factor (BDNF) or NT-4/5 in the brain tissues surrounding melanoma lesions. These studies support a role for NT and their receptors in the progression of melanomas to the brain-metastatic phenotype.