Parkin, PINK1 and DJ1 as possible modulators of mTOR pathway in ganglioglioma.

PURPOSE: Ganglioglioma (GG) is a non-malignant tumor classified as G1 by the WHO. Although we currently know that the neoplasm may result from the hyperactivity of protein kinase B (PKB or Akt) or extracellular-regulated kinase (Erk), which upregulates mammalian target of rapamycin kinase (mTOR) and leads to translation of proteins responsible for cell cycle regulation, there are still many questions to be answered. In the current paper we try to analyze the link between GG formation and activity of three proteins known to play a role in neuroprotection (parkin, PINK1 and DJ1). MATERIALS AND METHODS: In our paper, we review the current information on the involvement of these proteins in the transmission of information in the cell and triggering various cell signals, like survival or apoptosis. We also review current literature data on involvement of parkin, DJ1 and PINK1 in the regulation of mTOR, the pathway probably contributing to the development of GG. RESULTS: Parkin is an E3 ubiquitin ligase, shown to trigger proteasome-dependent degradation and autophagy, necessary for the maintenance of homeostasis in neurons. PINK1, a mitochondrial protein kinase, is required for mitochondrial maintenance and neuronal survival. DJ1 is a sensor of reactive oxygen species, and protects the cells against oxidative stress. Mutations in the genes encoding these three proteins are known to underlie autosomal recessive forms of Parkinson's disease, as well as other neurodegenerative and neuroinflammatory disorders. CONCLUSION: It appears that mutations of parkin, PINK1 and DJ1 may result in the development of both neurodegeneration and tumors. Also, these proteins might be used as markers of disease, thus allowing better diagnosis and therapy.

Focal cortical dysplasia: Molecular disturbances and clinicopathological classification (Review).

Focal cortical dysplasia (FCD) is one of the most important causes of drug-resistant epilepsy in paediatric patients, particularly in those below the age of 3. Even though over 40 years have passed since the first description of the entity by Taylor, the exact mechanisms causing these cortical abnormalities remain unelucidated. In this review, we summarise the current knowledge on clinical and histopathological aspects, taking into account the new classification system proposed by the International League Against Epilepsy. We focus on the clinicopathological associations and differences in post-surgical outcome among FCD subtypes, in particular isolated FCD vs. FCD associated with principal lesions, which have not been summarised to date. We also recapitulate genetic studies, pointing to the possible mechanisms of the cortical dysregulation and drug resistance, and summarise novel factors which may contribute to epileptogenesis in FCD. Furthermore, we compare FCD type IIB (FCDIIB) with brain tumours found in a neurocutaneous disorder, tuberous sclerosis, as we evaluate the hypothesis that FCD IIB may be a local form of this disease.

Medulloblastoma: molecular pathways and histopathological classification.

Malignant brain tumors are the leading cause of cancer death among pediatric patients, and medulloblastoma constitutes 20% of them. Currently, the treatment is risk-adapted. Maximum surgical resection is recommended, always followed by chemotherapy and neuroaxis radiotherapy. In spite of the improving survival rate, survivors succumb to treatment-induced side effects. To reduce toxic effects, molecular-targeted treatment is proposed. Medulloblastoma research is very robust, and new articles on the subject are published daily. In the current review we have tried to bring together molecular pathophysiology of the neoplasm and current pathological classification, thus making an effort to relate tumor biology and the histological picture.

Reactive oxygen species and synthetic antioxidants as angiogenesis modulators: Clinical implications.

Angiogenesis is important for normal functioning of organism and its disturbances are observed in many diseases, called angiogenesis-related states. Reactive oxygen species (ROSs) play an important role in physiology, but high level of cellular ROSs is cytotoxic and mutagenic for the cells, i.e. it can lead to oxidative stress. In this review we discuss close relationship between ROSs and angiogenesis process. Substances counteracting free radicals or their action and oxidative stress are known as antioxidants. We postulate that antioxidants, by affecting angiogenesis, may modulate therapy results in the case of angiogenesis-related disease. Herein, we present some antioxidant preparations of synthetic (N-acetylcysteine, curcumin and its analogs, Probucol, oleane tripertenoid, EGCG synthetic analogs) and nature-identical (vitamin E and C) origin. Then, we analyze their angiogenic properties and their multidirectional molecular effect on angiogenesis. Most preparations reduce neovascularization and diminish the level of proangiogenic molecules, downregulating signaling pathways related to angiogenesis. Moreover, we discuss studies concerning anticancer properties of presented synthetic antioxidants and their application in several angiogenesis-related diseases. We conclude that therapy in angiogenesis-related diseases should be planned with consideration of the angiogenic status of the patient.

Angiomodulatory properties of Rhodiola spp. and other natural antioxidants.

Disturbances of angiogenesis and oxidative stress can lead to many serious diseases such as cancer, diabetes or ischemic heart disease. Substances neutralizing oxidative stress are known as antioxidants. They can affect angiogenesis process also, and thus, they modulate therapy results. Antioxidants become more and more frequently used in order to maintain homeostasis of the organism and diminish the risk of disease. Herein, we introduce some antioxidant preparations of natural plant origin (Rhodiola, Aloe vera, Resveratrol, Echinacea, Plumbagin) and antioxidant supplements (Padma 28, Reumaherb, Resvega). Analyses of their angiogenic properties, their multidirectional molecular effect on angiogenesis as well as medical application are within the scope of this review. Most of presented preparations down regulate neovascularization. They can be safely administered to patients with abnormally high angiogenesis. Rhodiola modulates, and Echinacea, Aloe vera and Plumbagin inhibit tumour-related angiogenesis in vitro and in vivo (animal models). Resveratrol and Resvega reduce neovascularization in the eye and may be applicable in eye disorders. Padma 28 preparation exhibits angioregulatory activity, decreasing high angiogenesis of cancer cells and increasing physiological angiogenesis, therefore can be used in therapy of patients with various disturbances of angiogenesis. Antioxidant application in the case of angiogenesis-related diseases should take into consideration angiogenic status of the patient.

Pilocytic astrocytoma: a review of genetic and molecular factors, diagnostic and prognostic markers.

In spite of numerous studies concerning the pathogenesis of pilocytic astrocytoma (PA), the exact mechanism of the process still remains undetermined. It is difficult to obtain concordance between particular studies, which makes review of existing data especially troublesome. Nevertheless, the most important causative factors seem to be NF1 gene inactivation, in cases related to neurofibromatosis type 1, and BRAF gene overexpression in sporadic PAs, both resulting in MAPK/Erk pathway upregulation. Other molecular alterations, like mTOR or PI3K pathway deregulation, or Matrilin 2 overexpression, may influence the course of the disease, leading to the development of more aggressive and recurrent tumors. In the current paper we review genetic alterations in PA and describe currently studied molecular markers that may contribute to the development of the tumor and can be used in pathological staging of the malformation.

Upregulation of mitogen-activated protein kinase in ganglioglioma.

Ganglioglioma (GG) is a low-grade neoplasm, often associated with intractable epilepsy in pediatric patients. Available data suggest a relationship between GG and other glioneuronal lesions. So far, little is known about activation of kinases belonging to the mTor (mammalian target of rapamycin) pathway, although its upregulation is often found in brain tumors. Involvement of mTor kinase is responsible for excessive proliferation. In the current study we focused on the possible role of the Erk/Mapk (extracellular-signal-regulated kinase/mitogen-activated protein kinase) pathway in GG development. Eight GG tumors were resected from pediatric patients. Collected data reveal activation of proteins from the Erk pathway: Mek (extracellular regulated protein kinase kinase/mitogen-activated protein kinase kinase), Erk, Rsk1 (ribosomal S6 kinase 1), Rheb (Ras homolog enriched in brain) and Msk1 (mitogen- and stress-activated protein kinase 1), as detected by the western blot method. Moreover, activation of other proteins upstream of mTor - IGF-1R ß (insulin-like growth factor 1ß receptor), INR ß (insulin receptor ß), Akt/PKB (protein kinase B) - or downstream - 4E-BP1 (eukaryotic translation initiation factor 4E-binding protein 1) and rpS6 (ribosomal protein S6) - was also confirmed.

Frequency of TSC1 and TSC2 mutations in American, British, Polish and Taiwanese populations.

Tuberous sclerosis (TS) is caused by mutation of the tumor suppressor genes, tuberous sclerosis complex 1 (TSC1) or 2 (TSC2). The aim of the present study was to compare the frequency and types of TSC1 and TSC2 mutations in American, British, Polish and Taiwanese populations. A meta­analysis of 380 TS patients was performed. Significant differences were analyzed using the Chi-square test and one-way ANOVA analysis. Results showed a difference in frequency for the four populations analyzed. The frequency of TSC1 mutations was twice as high in the American and British populations. However, there were no significant differences in the types of mutations, with insertions of >1 nucleotide being the least frequent. Additionally, in an analysis of the complexity of nucleotide sequences it was demonstrated that the level of sequence complexity in the Polish population was significant higher compared to the remaining populations. Concerning strand bias, in the case of two types of substitutions, C>G/G>C and C>T/G>A, the ratio of corresponding mutations on the two DNA strands was approximately 3:1 and 2:1. In the present study, an increased frequency of C>G/G>C and C>T/G>A mutations in the coding strand was found in the analyzed populations. However, additional studies and larger patient cohorts are required to verify these results.

Fluvastatin influences hair color in C57BL/6 mice.

Our recent in vitro experiments suggest that fluvastatin may influence tyrosinase (key enzyme of melanogenesis) synthesis. The aim of the present study was to verify those findings in experiments, in vitro, in melanoma cell line, and in vivo, in mice. The expression of tyrosinase in B16F10 melanoma cell line, after induction of melanogenesis by UVB irradiation, was examined by Western blot analysis. Afterwards, the effect of fluvastatin on melanin synthesis in hair follicles of C57Bl/6 mice was investigated. The expression of tyrosinase was reduced in the presence of fluvastatin. In mice after anagen induction over the dorsal skin, gel containing fluvastatin in various concentrations was injected subcutaneously, while in part of control groups of mice, gel with placebo was injected. In addition, gel with fluvastatin was injected to four week-old mice (mice in first postnatal anagen) without anagen induction. In extension, injections of gel with fluvastatin or placebo were performed in mice without anagen induction (but after first postnatal anagen). In part of study group of mice (mice after anagen induction and injection of fluvastatin) regrowth of depigmented hair was observed, while in all control groups (mice after injection of placebo), such hair depigmentation over the skin area was not found. In conclusion, this study, for the first time, shows that fluvastatin might affect melanin synthesis in vivo.

Potential role of metalloproteinase inhibitors from radiation­sterilized amnion dressings in the healing of venous leg ulcers.

Chronic wounds are a significant socio-economic problem, thus, the improvement of the effectiveness of their treatment is an important objective for public health strategies. The predominant stage of the chronic wound is the inflammatory reaction which is associated with the damage of tissues, possibly due to the excessive secretion and activation of matrix metalloproteinases (MMPs). Several reports have suggested that amnion dressing inhibits tissue destruction and accelerates wound healing. Our recent study revealed that sterilized amnion stimulates keratinocyte proliferation in vitro, while the present study focused on the clinical application of radiation-sterilized amnion in chronic venous leg ulcers and aimed to explain the possible mechanism of its in vivo action. The study involved 25 individuals suffering from venous leg ulceration with a surface area of 10-100 cm2 and a healing rate below 10% per week, as verified during a 2-week screening period. The effectiveness of the amnion dressing was estimated following 4 weeks of treatment. The wound assessment, based on a modified Bates-Jensen Questionnaire, revealed a good and satisfactory response to the treatment in 23 of the 25 patients. The measurement of MMP-2 and MMP-9 activities in wound exudates revealed a decrease in activity in response to amnion application. This effect resulted from the presence of the potent MMP inhibitors, tissue inhibitor of metalloproteinases-1 (TIMP-1), type-1 plasminogen activator inhibitor (PAI-1) and thrombospondin-1 (TSP-1) in the amnion dressings, as shown by real-time fluorescence zymography and protein microarrays. Thus, unlike modern synthetic dressing materials, radiation-sterilized amnion dressings may have a multidirectional beneficial effect on chronic wounds.

Favourable prognosis in medulloblastoma with extensive nodularity is associated with mitogen-activated protein kinase upregulation.

Medulloblastoma with extensive nodularity (MBEN) is the only type of medulloblastoma (MB), an aggressive CNS tumour of childhood, that is connected with favourable prognosis. In patients with MBEN tumour resection and chemotherapy are sometimes sufficient. While development of other types of MB is usually connected with activation of the wingless pathway, sonic hedgehog pathway or mammalian target of rapamycin (mTor) pathway, little is known about the molecular basis of MBEN pathophysiology. In the present paper we evaluated activation of the mTor pathway and kinases upstream of mTor, mitogen-activated protein kinase (MAPK/Erk) and protein kinase B (PKB/Akt) in an MBEN sample. Using western blot technique with antibodies directed against active, phosphorylated forms of proteins, we found upregulation of mTor, Akt and Erk. Thus we postulate that the mTor pathway, often implicated in the development of CNS tumours, is also responsible for MBEN progression. Especially interesting seems implication of Erk and other kinases belonging to the same pathway: mitogen-activated protein kinase kinase (MEK-1) or phospho-ribosomal S6 kinase-1 (p90 RSK1), whose activity we usually demonstrate in more benign neoplasms. However, it remains to be clarified whether Erk pathway activation is actually prognostic for benign tumour development.

Activation of Akt/mTOR pathway in a patient with atypical teratoid/rhabdoid tumor.

A typical teratoid/rhabdoid tumor (AT/RT) is a highly malignant childhood brain tumor. Most AT/RTs are shown to contain chromosome 22 mutation in the region of hSNF5/INI1 gene, whose protein product participates in chromatin remodeling. Although the presence of this mutation is well described, molecular pathways underlying AT/RT development are poorly, if at all, understood. In the current paper we evaluate a case of AT/RT with special consideration of two pathways often implicated in tumor development: protein kinase B (PKB or Akt) and extracellular signal-regulated kinase (Erk). First, we confirmed expression of typical protein pattern being unique for AT/RT, including epithelial membrane antigen, S-100 and glial fibrillary acidic protein. In molecular analyses we tested the sample for activity of Akt and Erk kinase cascade. We found that Erk pathway signaling in the tumor was not upregulated. Neither c-Raf, MAPK nor Erk were hyperphosphorylated. On the other hand, we detected significant phosphorylation of Akt, phosphoinositide-dependent kinase-1 (PDK1) and glycogen synthase kinase 3 (GSK-3). At the same time, inhibitor of Akt pathway, phosphatase and tensin homolog (PTEN), was not upregulated. These results strongly support the hypothesis that Akt pathway contributes to chromatin remodeling disruption, promoting malignant transformation of AT/RT.

Fluvastatin increases tyrosinase synthesis induced by alpha-melanocyte-stimulating hormone in B16F10 melanoma cells.

The aim of this study was to evaluate the effect of fluvastatin on the alpha-melanocyte-stimulating hormone-mediated increase in tyrosinase activity in the melanoma B16F10 cell line and to establish whether Akt and extracellular signal-regulated kinase (Erk) inhibition is involved in tyrosinase synthesis after fluvastatin administration. Fluvastatin modulates alpha-melanocyte-stimulating hormone induced melanogenesis by increasing tyrosinase mRNA production, as shown by real time PCR, or tyrosinase protein synthesis, as presented by western blot technique. The stimulatory effect of fluvastatin on melanogenesis was, in part, induced by modulation of cell proliferation (decreased melanoma cell proliferation in G2/M phase) and possibly decrease of Akt. These findings indicate that fluvastatin increases tyrosinase synthesis induced by alpha-melanocyte-stimulating hormone in B16F10 cells and reveal an unknown effect of statin use: their influence on melanin production.

Occurrence of Ureaplasma parvum and Ureaplasma urealyticum in women with cervical dysplasia in Katowice, Poland.

The aim of this study was to evaluate the occurrence of genital mycoplasmas, especially Ureaplasma parvum and Ureaplasma urealyticum, in women with atypical squamous cells of undetermined significance (ASCUS), low grade squamous intraepithelial lesions (LSIL) and high grade squamous intraepithelial lesions (HSIL), compared to women with normal cytology living in Katowice, Poland. Two sterile swabs were used to obtain material from the posterior vaginal fornix of 143 women with squamous intraepithelial lesions and 39 healthy women: first for general bacteriology, second for detection of urogenital mycoplasmas using Mycoplasma IST2 kit. From each positive Mycoplasma IST2 culture DNA was isolated and PCR was performed for identification of U. parvum and U. urealyticum. Mycoplasma IST was positive in 34.1% cases. Urogenital mycoplasmas were demonstrated in women with HSIL significantly more often compared to women with LSIL, ASCUS, and with normal cytology. DNA of U. parvum was demonstrated in majority of Mycoplasma IST2-positive cases, U. urealyticum DNA-only in 9 (4.9%). Predominance of 3/14 serovars of U. parvum was demonstrated. U. urealyticum biovar 2 was present more often in women with squamous intraepithelial lesions.

Prevalence of urogenital mycoplasmas and ureaplasmas in women after kidney transplantation.

BACKGROUND: The prevalence of urogenital mycoplasmas and ureaplasmas in kidney transplant and hemodialyzed patients was studied. METHODS: Vaginal and cervical swabs taken from 40 women of the study group and 40 women of the control group were investigated. Identification of ureaplasmas, Mycoplasma genitalium, and human papillomavirus was performed by polymerase chain reaction. Each vaginal slide was evaluated for bacterial vaginosis. RESULTS: Urogenital mycoplasmas and ureaplasmas were significantly more common in patients from the study group (40%) compared with the control group (27.5%). Mycoplasma hominis and M. genitalium were demonstrated only in a few cases. Ureaplasma parvum was isolated predominantly, but Ureaplasma urealyticum was more common in patients from study group (10%) compared with control group (2.5%). In all U. urealyticum-positive women from the study group, human papillomavirus DNA was detected. CONCLUSION: Our observation showed the necessity of careful examination of possible atypical pathogens in diagnostic materials from hemodialyzed and kidney transplant patients.

Fibroblasts from normal skin of a tuberous sclerosis patient show upregulation of mTOR pathway.

Skin lesions are one of the characteristic features in tuberous sclerosis (TS), a neurocutaneous disorder caused by mutation of 1 of 2 tumor suppressor genes, encoding hamartin and tuberin. So far, however, studies on skin abnormalities present in TS patients are very few and do not contribute to the knowledge of the disease. In our current work, we cultured fibroblasts from healthy skin of a TS patient and evaluated upregulation of pathways found to be implicated in progression of TS tumors. We found that even healthy skin fibroblasts show upregulation of S6 ribosomal protein.

Cardiac rhabdomyomas in tuberous sclerosis complex show apoptosis regulation and mTOR pathway abnormalities.

Cardiac rhabdomyoma (CR) is the most common heart tumor in children and is usually associated with tuberous sclerosis complex (TSC). Tuberous sclerosis complex is a genetic disorder caused by a mutation in either of 2 genes (TSC1 or TSC2) and characterized by the formation of hamartomas in multiple organs. The 2 TSC proteins, hamartin and tuberin, antagonize the mammalian target of rapamycin (mTOR) signaling pathway, thus regulating cell growth and proliferation. Recently, some trials treating TSC with the mTOR inhibitor rapamycin have been published; however, the impact of such treatment on heart tumors is not known. The aim of the present paper was to study the molecular pathobiology of CRs. Six CR samples were studied. The expression of S6K1, pErk, Erk, Akt, pAkt, 4E-BP1, hamartin, tuberin, mTOR, bcl-2, Bax, and Ki-67 was examined using immunohistochemistry and Western blot methods. Increased expression of Bax, mTOR, pS6K, pErk, and 4E-BP1 was found in all CR samples. Hamartin and tuberin expression was decreased in tumors versus normal heart tissues. This is the first study showing mTOR pathway dysregulation and an increased expression of proapoptotic Bax protein in CRs associated with TSC.

Cardiac rhabdomyoma in tuberous sclerosis: hyperactive Erk signaling.

Tuberous sclerosis (TS) is a neurological disorder associated with the formation of tumors in several organs. Cardiac rhabdomyomas are possibly the earliest symptom of TS. Although rhabdomyomas are present in about half of TS patients, little is known of their molecular background since these tumors are rarely resected. Here we present a patient diagnosed with TS, in whom rhabdomyoma has been excised due to deterioration of hemodynamics. We found, that the tumor remained heterozygous for the affected TSC2 gene. To analyze molecular mechanisms implicated in rhabdomyoma growth, we determined the status of mTOR, Akt and Erk pathways. We found that Akt was not upregulated, while mTOR, Erk and its substrates were hyperactive. Classic activator of Erk, MEK, was only modestly active. We hypothesize that rhabdomyoma arising in TS may progress due to Erk potentiation.

Relationship between serum progesterone and tumor necrosis factor production in postmenopausal women undergoing estrogen/medroxyprogesterone therapy.

Changes in tumor necrosis factor (TNF) production by lipopolysaccharide-stimulated peripheral blood mononuclear cells significantly correlated with serum P levels in postmenopausal women subjected to estrogen/medroxyprogesterone therapy. This may suggest that low physiologic or hormone therapy-related changes of serum P in healthy postmenopausal women may affect an ability of peripheral blood mononuclear leukocytes to produce TNF, thus having an impact on a variety of TNF-dependent physiologic and pathologic phenomena.

Fluvastatin increases tyrosinase synthesis induced by UVB irradiation of B16F10 melanoma cells.

Statins are widely used to lower plasma concentrations of lipids, e.g. cholesterol. One of the main effects of statin treatment is inhibition of hydroxymethyl glutaryl-coenzyme A reductase. The role of fluvastatin, a frequently used statin, was examined in potential modulation of tyrosinase (key enzyme of melanogenesis) synthesis. Levels of tyrosinase mRNA induced by UVB irradiation of B16F10 melanoma cell line were measured by real time PCR. Fluvastatin increases tyrosinase mRNA production induced by UVB irradiation in B16F10 melanoma cell line. Fluvastatin treatment may potentially influence melanin synthesis and protection against UV irradiation.