Antidepressant-like effect of endomorphin-1 and endomorphin-2 in mice.

Endomorphin-1 (Tyr-Pro-Trp-Phe-NH(2)) and endomorphin-2 (Tyr-Pro-Phe-Phe-NH(2)) are two recently isolated mu-opioid selective peptides with a potent antinociceptive activity, involved in a number of physiological processes, including food intake, vasomotricity, sexual behavior, as well as neuroendocrine and cardiorespiratory functions. The neuroanatomical distribution of endomorphins prompted us to study their antidepressant activity in two animal behavioral models of depression: forced-swimming and tail-suspension tests. In both tests, the intracerebroventricular (i.c.v.) injection of either endomorphin-1 or endomorphin-2 significantly decreased the duration of immobility, interpreted as an expression of 'behavioral despair', which could be related to the depression syndrome. These effects of endomorphins did not result from the stimulation of the animal motor activity. We have also demonstrated that the antidepressant-like effect of endomorphins was antagonized by the universal opioid antagonist, naloxone and the mu-opioid receptor selective antagonist, beta-funaltrexamine. In contrast, this effect was not antagonized by delta- and kappa-opioid receptor selective antagonists, naltrindole and nor-binaltorphimine, respectively. The results of the present study demonstrate that endomorphin-1 and endomorphin-2 produce potent antidepressant-like effects after i.c.v. injection in mice. We may suggest that endomorphins and the mu-opioid receptors might be involved in the physiopathology of depressive disorders, and that the endomorphinergic system could serve as a novel target for the development of antidepressant drugs.     

The endomorphin system and its evolving neurophysiological role

Endomorphin-1 (Tyr-Pro-Trp-Phe-NH2) and endomorphin-2 (Tyr-Pro-Phe-Phe-NH2) are two endogenous opioid peptides with high affinity and remarkable selectivity for the mu-opioid receptor. The neuroanatomical distribution of endomorphins reflects their potential endogenous role in many major physiological processes, which include perception of pain, responses related to stress, and complex functions such as reward, arousal, and vigilance, as well as autonomic, cognitive, neuroendocrine, and limbic homeostasis. In this review we discuss the biological effects of endomorphin-1 and endomorphin-2 in relation to their distribution in the central and peripheral nervous systems. We describe the relationship between these two mu-opioid receptor-selective peptides and endogenous neurohormones and neurotransmitters. We also evaluate the role of endomorphins from the physiological point of view and report selectively on the most important findings in their pharmacology.     

Ciglitazone, an agonist of peroxisome proliferator-activated receptor gamma, exerts potentiated cytostatic/cytotoxic effects against tumor cells when combined with lovastatin

Thiazolidinediones are ligands of PPAR-gamma, a member of the nuclear receptor family. These drugs have shown promising pre-clinical activity in tumor models but clinical studies failed to confirm their beneficial effect. We have studied the in vitro antitumor effects of a combination of ciglitazone, a thiazolidinedione drug, and lovastatin, an inhibitor of 3-hydroxy-3-methylglutaryl-coenzyme A reductase. We observed a marked synergism in several different tumor cell lines resulting from both inhibition of cell proliferation and induction of apoptosis. These results strongly suggest that combining PPAR-gamma agonists with statins can produce significant antitumor effects.     

Nestin as a marker of cancer stem cells

The crucial role of cancer stem cells (CSCs) in the pathology of malignant diseases has been extensively studied during the last decade. Nestin, a class VI intermediate filament protein, was originally detected in neural stem cells during development. Its expression has also been reported in different tissues under various pathological conditions. Specifically, nestin has been shown to be expressed in transformed cells of various human malignancies, and a correlation between its expression and the clinical course of some diseases has been proved. Furthermore, the coexpression of nestin with other stem cell markers was described as a CSC phenotype that was subsequently verified using tumorigenicity assays. The primary aim of this review is to summarize the recent findings regarding nestin expression in CSCs, its possible role in CSC phenotypes, particularly with respect to capacity for self‐renewal, and its utility as a putative marker of CSCs.     

Adeno-associated virus-mediated delivery of a mutant endostatin in combination with carboplatin treatment inhibits orthotopic growth of ovarian cancer and improves long-term survival

A human ovarian cancer cell line, which migrates to mouse ovaries and establishes peritoneal carcinomatosis, was used to evaluate the cooperative effect of an antiangiogenic gene therapy combined with chemotherapy. The ovarian carcinoma cell line MA148 was genetically modified by "Sleeping Beauty" transposon-mediated delivery of DsRed2 fluorescent protein. Stable, high-level expression of DsRed protein enabled in vivo imaging of peritoneal dissemination of ovarian cancer. Both external and internal imaging, along with histopathology, showed migration of i.p. injected human ovarian cancer cell line to mouse ovaries. Using this model, we evaluated the effect of adeno-associated virus (AAV)-mediated expression of a mutant endostatin either alone or in combination with carboplatin treatment. A single i.m. injection of recombinant AAV (rAAV)-mutant human endostatin with P125A substitution (P125A-endostatin) showed sustained expression of mutant endostatin. Antiangiogenic gene therapy inhibited orthotopic growth of ovarian cancer and resulted in 33% long-term tumor-free survival. A single cycle of carboplatin treatment combined with mutant endostatin gene therapy resulted in 60% of the animals remaining tumor free for >200 days, which was significantly better than rAAV-LacZ and/or carboplatin. Combination treatment delayed tumor appearance in 40% of the animals, wherein the residual tumors were smaller in size with limited or no peritoneal metastasis. These studies suggest that AAV-mediated gene therapy of P125A-endostatin in combination with carboplatin is a useful method to inhibit peritoneal dissemination of ovarian carcinoma.     

The role of actin in the apoptotic cell death of P19 embryonal carcinoma cells

The P19 mouse embryonal carcinoma cell line was used as a model for a study of apoptosis accompanying differentiation induced by all-trans retinoic acid (ATRA). Apoptosis was detected both on the basis of morphological features (nuclear fragmentation, blebbing of plasma membrane, and formation of apoptotic bodies), and by using DNA electrophoresis and flow-cytometric measurement of DNA content. Actin cytoskeleton was studied both on morphological and submicroscopic levels. ATRA-treated cells manifested apoptosis-specific changes in the distribution of actin foremost in association with their entry into executive phase of apoptosis, when F-actin cables participated in cell disintegration into apoptotic bodies. Using immunogold labeling, actin was also identified in centers of fragmenting apoptotic nuclei, in the disintegration of which it is likely involved as well. At the same time, a cleavage of actin by active caspase-3 was proved, resulting in the emergence of 32 kDa fragment, termed fractin. Measurement of F-actin and fractin content using flow cytometry showed an unequivocal decrease of F-actin and synchronous increase of fractin in the apoptotic population as compared to non-treated cells. Therefore, our results proved both actin proteolysis and active involvement of specific actin structures in the final cell disintegration during apoptosis in the P19 cells.     

Cathepsin D expression in human colorectal cancer: relationship with tumour type and tissue differentiation grade

OBJECTIVE: Cathepsin D (CD) is one of the main proteolytic enzymes contributing to the development of cancer. The aim of this study was to CD activity assay in the homogenates of tissues from the centre of the tumour (0) and tumour free area 2 cm, and 5 cm from the tumour border in human colorectal cancer. Activity in the centre of the tumour was compared with immunohistochemical expression CD. METHODS: CD activity was measured using acid denatured Hb as a substrate. For immunohistochemical staining peroxidase method was used. RESULTS: Activity of CD was significantly higher (15-fold) in tumour tissue homogenates in comparison to normal mucosa adjacent (control) (p < or = 0.0001) and raised parallel to the stage of tumour tissue differentiation grade. CD activity decreased significantly (p < or = 0.0001) with the distance from the tumour border 2 cm (12.7 fold) and 5 cm (5.7 fold) in comparison to the centre of the tumour. In immunohistochemical examinations CD was detected as diffuse cytoplasmic as well as fine granular staining of the cytoplasm, with occasional coarse cytoplasmic granules staining in the same cases that were positive for both. Positive staining was observed in 2 of 3 in well-differentiated (66%), 4 of 10 in moderately-differentiated (40%) and 4 of 5 in poorly-differentiated (80%), tubular adencarcinomas represented: 3 of 7 (42%) and 9 of 13 in invasive adencarcinoma (69%). CONCLUSION: We have observed a wide range of cathepsin D and their antigen expressions patterns in colorectal tumours with the development the disease stage, this finding may be used as a daignostic tumor marker in colorectal cancer.     

The effect of chemotherapy on status of estrogen receptors in primary tumors and lymph node metastases of human ductal breast cancer

The aim of the study was the evaluation of ERalpha and ERbeta expression in primary tumors and lymph node metastases of breast cancer as well as the assessment of the influence of preoperative chemotherapy on these receptors with regard to changes in morphological appearance of primary tumors and their metastases. Immunohistochemical examinations were conducted on surgically removed ductal invasive breast cancers and their lymph node metastases of 135 patients. Seventy-one patients were spared preoperative chemotherapy which was administered to other 64 patients. Primary breast cancers with preoperative chemotherapy showed lower mean percentage of cells with a positive reaction to ERalpha and ERbeta as compared to primary tumors without preoperative chemotherapy. There were positive correlations among primary tumors and lymph node metastases regardless of preoperative chemotherapy applied. On the other hand, ERalpha and ERbeta expressions were negatively correlated in primary tumors without chemotherapy in contrast to primary tumors after chemotherapy. Furthermore, it was observed that preoperative chemotherapy was responsible for significantly less damage to lymph node metastases of breast cancer in comparison to primary tumors. In cases of such advanced damage of primary tumors that made determination of estrogen receptor expression impossible, their evaluation was performed on metastases to regional lymph nodes. Although preoperative chemotherapy did not severely impair estrogen receptor expression, presented changes of their distribution are a sufficient reason for simultaneous labeling of estrogen receptors in both primary tumors and metastases due to various sensitivity to chemotherapy of primary cancers in comparison with involved lymph nodes.     

Association of axon guidance factor semaphorin 3A with poor outcome in pancreatic cancer

Neural alterations and aberrantly expressed nerve-specific factors promoting tumor progression are known to contribute to pancreatic cancer's extremely poor prognosis. Despite hints that axon guidance factor semaphorin 3A (SEMA3A) may function as a tumor inhibitor, its clinical importance and therapeutic potential have not yet been explored. The present study investigated the role of SEMA3A and its receptors-plexins A1-A4 (PLXNA1-A4) and neuropilin-1 (NRP1)-in pancreatic cancer. QRT-PCR and immunohistochemical analyses revealed overexpression of SEMA3A, NRP1 and PLXNA1 in metaplastic ducts, malignant cells and nerves of cancerous specimens, and showed that elevated levels of corresponding mRNA (6.8-fold, 2.0-fold and 1.5-fold, respectively) clearly correlated with negative clinicopathological manifestations such as shorter survival (SEMA3A and PLXNA1) and a lesser degree of tumor differentiation (NRP1) in Stages I-III patients. High SEMA3A expression in pancreata of Stage IV M1 patients and in peritoneal metastases, and consequent functional studies indicated that poor clinical outcome might be related to the ability of SEMA3A to promote dissemination and invasiveness of pancreatic cancer cells through activation of multiple pathways involving Rac1, GSK3b or p42/p44 MAPK, but not E- to N-cadherin switch, MMP-9 or VEGF induction. Thus, this study is the first to quantify expression of the SEMA3A system in human malignancy and to show that overexpression of SEMA3A by nerves and transformed cells leads to a SEMA3A-rich environment which may favor malignant activities of tumor cells. Furthermore, negative clinicopathological correlations suggest that SEMA3A might represent a novel intervention target but not a treatment option for pancreatic cancer patients.