Distribution of nerve endings in human thumb interphalangeal joint.

This study aims to quantitatively analyze the distribution of encapsulated nerve endings in the human thumb interphalangeal (IP) joint capsule. There are three types of nerve endings. Type-I nerve endings (Ruffini-like ending) sense pressure changes, Type II (Pacini-like ending) nerve endings contribute to the kinesthetic sense, and Type III (Golgi-like ending) nerve ending provides proprioceptive information. We dissected five right thumbs IP joints from freshly frozen cadavers (5 men). The mean age of the cadavers at the time of death was 63.4 years (55-73). Sections were stained with the hematoxylin-eosin and antiprotein gene product 9.5 (PGP9.5) to identify encapsulated nerve endings. Transverse sections were cut and divided into volar, dorsal, and then into two equal parts, proximal and distal. The density of encapsulated nerve endings compared to volar versus dorsal and proximal versus distal regions was examined. This study showed that type 1 nerve endings were more common in the distal parts of the IP joint (p < 0.05). Also, type 3 nerve endings were observed in the thumb IP joint. There was no difference between regions in type II and type III nerve endings. The current study demonstrates that the distribution of encapsulated nerve endings in the IP joint is different from the PIP and DIP joints. Moreover, further studies are required to understand the thumb's physiology.

SIRT1/FOXO Signaling Pathway in Breast Cancer Progression and Metastasis.

Breast cancer is the second most common cancer in women. The roles of the SIRT and FoxO proteins in tumor progression are known, but their roles in metastasis have not yet been clearly elucidated. In our study, we investigated the roles of SIRT and FoxO proteins their downstream pathways, proteins p21 and p53, in tumor progression and metastasis. We evaluated these proteins in vitro using metastatic 4TLM and 67NR cell lines, as well as their expression levels in tumor-bearing mice. In addition, the regulatory role of SIRT and FoxO proteins in different transduction cascades was examined by IPA core analysis, and clinicopathological evidence was investigated in the TCGA database. In primary tumors, the expression levels of SIRT1, p21, p53, E2F1 and FoxO proteins were higher in 67NR groups. In metastatic tissues, the expression levels of SIRT1, E2F1 and FoxO proteins were found to be enhanced, whereas the levels of p53 and p21 expression were noted to be reduced. IPA analysis also provided empirical evidence of the mechanistic involvement of SIRT and FoxO proteins in tumor progression and metastasis. In conclusion, SIRT1 was found to co-operate with FoxO proteins and to play a critical role in metastasis. Additional research is required to determine why overexpression of SIRT1 in metastatic tissues has oncogenic effects.

CD200 mimetic aptamer PEG-M49 markedly increases the therapeutic effects of pegylated liposomal doxorubicin in a mouse model of metastatic breast carcinoma: an effect independent of CD200 receptor 1.

We previously reported that CD200 overexpression in the host decreases progression and metastasis of the highly aggressive metastatic 4THM breast carcinoma. We have explored a possible synergistic interaction between the CD200 mimetic PEG-M49 and pegylated liposomal doxorubicin (Peg-Dox) in wild-type CD200 knockout (CD200-/-) and CD200 Receptor 1 knockout (CD200R1-/-) mice for the first time. A 4THM breast carcinoma model and three groups of BALB/c mice (wild type, CD200-/- and CD200R1-/-) were used. Five days after injection of tumor cells, mice were injected with Peg-Dox (ip, once a week) and PEG-M49 or a control aptamer (iv, every 3 days). Necropsies were performed either 12 (mid-point) or 24 (endpoint) days after injection and the extent of tumor growth, visceral metastasis and changes in the tumor-directed immune response were evaluated. PEG-M49 and Peg-Dox co-treatment induced complete tumor regression and loss of macroscopic lung metastasis in four out of seven WT mice. This synergistic anti-tumoral effect is thought to be due to Peg-M49-induced inhibition of Gr1 + CD11b + cells and Peg-Dox-induced increases in tumor-infiltrating CD8 + and CD8CD4 double-positive cells. Similar changes were observed in CD200R1-/- mice indicating that the primary effects of Peg-M49 are mediated by non-CD200R1 receptors. We also demonstrated for the first time that tumor growth, metastasis, and tumor infiltrating GR1 + CD11b + cells were markedly increased in CD200R1-/- mice, indicating an anti-inflammatory and protective role of CD200. CD200 mimetics might be a safe and effective immunomodulatory treatment in conjunction with classical chemotherapeutics for therapy of aggressive metastatic breast carcinoma.

Regulation of NLRP3 by non-coding RNAs in different cancers: interplay between non-coding RNAs and NLRP3 in carcinogenesis and metastasis.

NLRP3 (NOD-, LRR- and pyrin domain-containing protein 3) inflammasomes are multitasking intracellular sensors having characteristically unique ability to detect myriad of microbial motifs and endogenous danger signals which promote structural assembly of NLRP3 inflammasome thus enabling it to perform instrumental roles. Detailed mechanistic insights revealed that molecularly assembled NLRP3 inflammasomes stimulated caspase-1-driven release of the pro-inflammatory cytokines. NLRP3 has been shown to play fundamental role in the regulation of cancer progression and metastasis. Recently emerging cutting-edge research-works have started to shed light on the involvement of non-coding RNAs in the regulation of NLRP3 in different cancers. MicroRNAs, lncRNAs and circular RNAs have been shown to modulate NLRP3 in different diseases. However, we still have incomplete information about regulation of NLRP3 by circular RNAs in various cancers. In this review, we will comprehensively analyze how different microRNAs and long non-coding RNAs modulate NLRP3 in human cancers. Emerging evidence has started to scratch the surface of the participation of miRNAs and lncRNAs in the regulation of NLRP3. Xenografted mice-based studies have also enabled us to develop a better comprehension of interplay between miRNAs, lncRNAs and NLRP3. Hopefully, detailed analysis of contextual regulation of NLRP3 by oncogenic and tumor suppressor miRNAs, lncRNAs and circRNAs will be helpful in getting a step closer to the personalized medicine.

Central apelin mediates stress-induced gastrointestinal motor dysfunction in rats.

Apelin, an endogenous ligand for APJ receptor, has been reported to be upregulated in paraventricular nucleus (PVN) following stress. Central apelin is known to stimulate release of corticotropin-releasing factor (CRF) via APJ receptor. We tested the hypothesis that stress-induced gastrointestinal (GI) dysfunction is mediated by central apelin. We also assessed the effect of exogenous apelin on GI motility under nonstressed (NS) conditions in conscious rats. Prior to solid gastric emptying (GE) and colon transit (CT) measurements, APJ receptor antagonist F13A was centrally administered under NS conditions and following acute stress (AS), chronic homotypic stress (CHS), and chronic heterotypic stress (CHeS). Plasma corticosterone was assayed. Strain gage transducers were implanted on serosal surfaces of antrum and distal colon to record postprandial motility. Stress exposure induced coexpression of c-Fos and apelin in hypothalamic PVN. Enhanced hypothalamic apelin and CRF levels in microdialysates were detected following AS and CHeS, which were negatively and positively correlated with GE and CT, respectively. Central F13A administration abolished delayed GE and accelerated CT induced by AS and CHeS. Central apelin-13 administration increased the plasma corticosterone and inhibited GE and CT by attenuating antral and colonic contractions. The inhibitory effect elicited by apelin-13 was abolished by central pretreatment of CRF antagonist CRF9-41 in antrum, but not in distal colon. Central endogenous apelin mediates stress-induced changes in gastric and colonic motor functions through APJ receptor. The inhibitory effects of central exogenous apelin-13 on GI motility appear to be partly CRF dependent. Apelin-13 inhibits colon motor functions through a CRF-independent pathway.

Pentoxifylline Alleviates Early Brain Injury in a Rat Model of Subarachnoid Hemorrhage.

BACKGROUND: Subarachnoid hemorrhage (SAH) is a severe cerebrovascular disease frequently caused by ruptured aneurysms. Early brain injury (EBI) is the primary cause of morbidity and mortality in patients diagnosed with SAH and is associated with increased intracranial pressure, decreased cerebral blood flow and cerebral ischemia. Pentoxifylline (PTX) is a methylxanthine derivative clinically proven to improve perfusion in the peripheral microcirculation and has been shown to have neuroprotective effects in brain trauma and global cerebral ischemia in experimental animal models. This study aimed to determine the effect of PTX in experimental SAH, which has not been investigated yet. METHODS: An experimental SAH model was induced in male Wistar rats by autologous blood injection into the prechiasmatic cistern, and PTX was injected intraperitoneally immediately after SAH. The effects of PTX were evaluated 24 h after SAH via assessing the cerebral ultrastructure via transmission electron microscopy (TEM). Brain edema, blood-brain barrier (BBB) permeability, red blood cell deformability, tumor necrosis factor-alpha (TNF-alpha), nitrite-nitrate levels and apoptotic neuron death were also determined 24 h after SAH. The BBB permeability was measured by Evans blue (EB) extravasation, erythrocyte deformability was determined by filtration technique, and TNF-alpha and reactive nitrogen metobolites were analyzed in brain tissue by ELISA and spectral analysis, respectively. Apoptotic neurons were determined in brain sections by cleaved caspase-3 immunohistochemical analysis, and expression intensity was quantified using image J software. RESULTS: Cerebral ultrastructure in SAH group animals revealed intense perivascular edema and distortion in the astrocyte foot processes. PTX treatment attenuated structural deterioration due to SAH. Brain water content, BBB permeability, TNF-alpha, nitrite-nitrate levels and apoptotic neuronal death were significantly increased 24 h after SAH and were significantly alleviated by PTX treatment. There was no significant change in red cell deformability after SAH. CONCLUSIONS: Our results show that PTX reduces brain edema, BBB permeability, TNF-alpha expression, reactive nitrogen metobolites and apopotosis in experimental SAH. Based on our findings we suggest that PTX exerts neuroprotection against SAH-induced EBI, which might be associated with the inhibition of inflammation and apoptotic neuronal cell death.

Autocrine control of MIP-2 secretion from metastatic breast cancer cells is mediated by CXCR2: a mechanism for possible resistance to CXCR2 antagonists.

CXCR2 interacts with a wide range of chemokines and CXCR2 antagonists may have therapeutic value for treatment-resistant metastatic carcinomas. We aimed to explore regulation of activity of CXCR2 and its ligand, MIP-2, in metastatic breast carcinoma. We used mouse breast carcinoma cells metastasize to brain (4TBM), liver (4TLM), and heart (4THM) and explored the extra- and intracellular mechanisms effecting MIP-2 secretion using CXCR2 antagonist and inhibitors of downstream signaling molecules. 4TBM, 4TLM, and 4THM cells include cancer stem cell features and metastasize extensively. We also determined kinetics of MIP-2 secretion in 4T1 and non-metastatic 67NR mouse breast carcinoma cells. We found that there is an autocrine-inhibition of MIP-2 secretion. Specifically, metastatic cells selectively express CXCR2 only, and not CXCR1 and attenuating CXCR2 activity with SB225002 increased MIP-2 secretion. This may be due to the inhibition of protein kinase C (PKC) activity since RO318220; a specific inhibitor of PKC also increased MIP-2 secretion. Attenuating CXCR2 activity with SB225002, otherwise suppressed proliferation of 4THM and 4TBM cells. Tumor explants and cancer-associated fibroblasts obtained from 4TLM, 4THM, and 4TBM primary tumors secreted high levels of MIP-2. Surprisingly, CXCR2 expression was low in 4TLM cells demonstrating that liver metastatic cells might be resistant to the anti-tumoral effects of CXCR2 antagonists. Our results demonstrated that resistance to anti-proliferative effects of CXCR2 may also arise from feedback increases in MIP-2 secretion. Activation of PI3 K pathway augments MIP-2 secretion, hence possible resistance to the antitumor effects of CXCR2 antagonists might be prevented with inhibitors of PI3 K.

Activation of neuroimmune pathways increases therapeutic effects of radiotherapy on poorly differentiated breast carcinoma.

Recent studies document the importance of neuronal dysfunction in cancer development and metastasis. We reported previously that both depletion of neuropeptides in capsaicin-sensitive sensory nerve endings and vagotomy increases metastasis of triple negative breast carcinoma. Of the sensory neuropeptides, Substance P (SP) is distributed widely for regulation of immune functions. We therefore examined the affects of continuous exposure to low doses of SP on brain metastatic cells of the mouse breast carcinoma (4TBM) in the presence of radiotherapy (RT) thought to increase antigenicity of cancer cells. 4TBM cells have a cancer stem cell phenotype and induce extensive visceral metastasis after orthotopic inoculation into the mammary pad. Results demonstrated that SP treatment decreases the number of tumor-infiltrating myeloid-derived suppressor cells as well as the TNF-α response to LPS challenge. SP also increased CD4+Cd25(bright) cells in draining lymph nodes of tumor-bearing animals and IFN-γ secretion from leukocyte culture prepared from lymph nodes and spleens of tumor-bearing animals. SP also prevented tumor-induced degeneration of sensory nerve endings and altered release of angiogenic factors from cancer-associated fibroblasts (CAF) and tumor explants. In accordance with these observed immunological effects, combination treatment of continuous SP with a single dose of RT induced complete tumor regression and significantly reduced or prevented metastasis in 50% of the animals while suppressing primary tumor growth and metastasis in the remaining mice. These original findings demonstrate that SP through neuroimmune modulation can prevent formation of immune suppression in the tumor microenvironment, enhance cytotoxic immunity in the presence of RT and prevent metastatic growth.

Expression of CCM2 and CCM3 during mouse gonadogenesis.

PURPOSE: Three cerebral cavernous malformation (CCM) proteins, CCM1, CCM2, and CCM3, regulate cell-cell adhesion, cell shape and polarity, and most likely cell adhesion to extracellular matrix. Recently, CCM2 and CCM3 are known to be expressed in control and varicocele-induced rat testes, but little is known about these proteins during gonadogenesis. This led us to study the CCM proteins during the mouse gonadogenesis. METHODS: Neonatal (PND 0), postnatal, and adult mice testes and ovaries were obtained from mice. CCM2 and CCM3 expression were analyzed during mouse testicular and ovarian development by immunohistochemistry and quantitative real-time PCR. RESULTS: The results showed that in both sexes, Ccm2 and Ccm3 mRNA and protein were first detectable after gonadogenesis when the gonads were well differentiated and remained present until the adult stage. In the testis, CCM2 and CCM3 expression were restricted to the nuclei of Sertoli cells, suggesting a conserved role in testicular differentiation. In the ovary, the CCM2 and CCM3 proteins were localized in the cytoplasm of oocytes, suggesting an unexpected role during oogenesis. Quantitative real-time PCR (qRT-PCR) results showed that expression of Ccm2 and Ccm3 genes could play a role in the regulation of mouse gonadogenesis translational activation upon testicular and ovarian development. CONCLUSIONS: The localization of CCM2 and CCM3 proteins show their different functions for CCM2 and CCM3 which may have important roles in testicular and ovarian differentiation. In conclusion, CCM2 and CCM3 may be involved in establishing the differential expression pattern in developing mouse testis and ovary.

Hypertension alters phosphorylation of VASP in brain endothelial cells.

Hypertension impairs cerebral vascular function. Vasodilator-stimulated phosphoprotein (VASP) mediates active reorganization of the cytoskeleton via membrane ruffling, aggregation and tethering of actin filaments. VASP regulation of endothelial barrier function has been demonstrated by studies using VASP(-/-) animals under conditions associated with tissue hypoxia. We hypothesize that hypertension regulates VASP expression and/or phosphorylation in endothelial cells, thereby contributing to dysfunction in the cerebral vasculature. Because exercise has direct and indirect salutary effects on vascular systems that have been damaged by hypertension, we also investigated the effect of exercise on maintenance of VASP expression and/or phosphorylation. We used immunohistochemistry, Western blotting and immunocytochemistry to examine the effect of hypertension on VASP expression and phosphorylation in brain endothelial cells in normotensive [Wistar-Kyoto (WKY)] and spontaneously hypertensive (SH) rats under normal and exercise conditions. In addition, we analyzed VASP regulation in normoxia- and hypoxia-induced endothelial cells. Brain endothelial cells exhibited significantly lower VASP immunoreactivity and phosphorylation at the Ser157 residue in SHR versus WKY rats. Exercise reversed hypertension-induced alterations in VASP phosphorylation. Western blotting and immunocytochemistry indicated reduction in VASP phosphorylation in hypoxic versus normoxic endothelial cells. These results suggest that diminished VASP expression and/or Ser157 phosphorylation mediates endothelial changes associated with hypertension and exercise may normalize these changes, at least in part, by restoring VASP phosphorylation.

P/Q type (Cav2.1) Calcium Channel Blocker ω-Agatoxin IVA Alters Cleaved Caspase-3 and BDNF Expressions in the Rat Brain and Suppresses Seizure Activity.

High-voltage-gated calcium channels have pivot role in the cellular and molecular mechanisms of various neurological disorders, including epilepsy. Similar to other calcium channels, P/Q-type calcium channels (Cav2.1) are also responsible for vesicle release at synaptic terminals. Up to date, there are very limited reports showing the mechanisms of Cav2.1 in epileptogenesis. In the present study, we investigated the anticonvulsive and neuroprotective effects of ω-agatoxin IVA, a specific Cav2.1 blocker, in a chemical kindling model of epileptogenesis. Righting reflex and inclined plane tests were used to assess motor coordination. Electroencephalography was recorded for electrophysiological monitoring of seizure activity in freely moving rats. Immunohistochemical analyses were performed for brain-derived neurotrophic factor (BDNF) and cleaved caspase-3 expressions in the prefrontal cortex, striatum, hippocampus, and thalamic nucleus. ω-Agatoxin IVA injected into the right lateral ventricle significantly prolonged the onset of seizures in a dose-dependent manner. In addition, repeated intraperitoneal administrations of ω-agatoxin IVA significantly suppressed the development of kindling and epileptic discharges without altering motor coordination. In addition, ω-agatoxin IVA significantly increased BDNF expressions, and decreased cleaved caspase-3 expressions in the brain when compared to PTZ + saline group. Our current study emphasizes the significance of the inhibition of P/Q type calcium channels by ω-agatoxin IVA, which suppresses the development of epileptogenesis and provides a new potential pathway for epilepsy treatment.

Cerebral cavernous malformation proteins, CCM1, CCM2 and CCM3, are decreased in metastatic lesions in a murine breast carcinoma model.

Three genes are associated with cerebral cavernous malformations (CCMs): CCM1, CCM2 and CCM3. These genes participate in microvascular angiogenesis, cell-to-cell junctions, migration and apoptosis. We evaluated the expression in vivo of CCM genes in primary tumors and metastastases in a murine model of metastatic breast carcinoma. We used cell lines obtained from metastasis of 4T1, 4TLM and 4THM breast cancer to liver and heart. These cells were injected into the mammary ridge of Balb/C female mice. After 27 days, the primary tumors, liver and lung were removed and CCM proteins were assessed using immunohistochemistry and western blot analysis. CCM proteins were expressed in primary tumor tissues of all tumor-injected animals; however, no CCM protein was expressed in metastatic tumor cells that migrated into other tissues. CCM proteins still were observed in the lung and liver tissue cells. Our findings suggest that CCM proteins are present during primary tumor formation, but when these cells develop metastatic potential, they lose CCM protein expression. CCM protein expression was lost or reduced in metastatic tissues compared to the primary tumor, which indicates that CCM proteins might participate in tumorigenesis and metastasis.

Protective Effects of Adropin in Experimental Subarachnoid Hemorrhage.

PURPOSE: We aimed to investigate early effects of exogenously administered adropin (AD) on neurological function, endothelial nitric oxide synthase (eNOS) expression, nitrite/nitrate levels, oxidative stress, and apoptosis in subarachnoid hemorrhage (SAH). METHODS: Following intracerebroventricular AD administration (10 µg/5 µl at a rate of 1 µl/min) SAH model was carried out in Sprague-Dawley rats by injection of autologous blood into the prechiasmatic cistern. The effects of AD were assessed 24 h following SAH. The modified Garcia score was employed to evaluate functional insufficiencies. Adropin and caspase-3 proteins were measured by ELISA, while nitrite/nitrate levels, total antioxidant capacity (TAC) and reactive oxygen/nitrogen species (ROS/RNS) were assayed by standard kits. eNOS expression and apoptotic neurons were detected by immunohistochemical analysis. RESULTS: The SAH group performed notably lower on the modified Garcia score compared to sham and SAH + AD groups. Adropin administration increased brain eNOS expression, nitrite/nitrate and AD levels compared to SHAM and SAH groups. SAH produced enhanced ROS/RNS generation and reduced antioxidant capacity in the brain. Adropin boosted brain TAC and diminished ROS/RNS production in SAH rats and no considerable change amongst SHAM and SAH + AD groups were detected. Apoptotic cells were notably increased in intensity and number after SAH and were reduced by AD administration. CONCLUSIONS: Adropin increases eNOS expression and reduces neurobehavioral deficits, oxidative stress, and apoptotic cell death in SAH model. Presented results indicate that AD provides protection in early brain injury associated with SAH.

Differential characteristics of heart, liver, and brain metastatic subsets of murine breast carcinoma.

Breast carcinoma is comprised of heterogeneous groups of cells with different metastatic potential. To develop effective therapeutic strategies targeting metastatic disease, it is crucial to understand the characteristics of breast cancer cells that enable metastasis to distant organs. 4THM breast carcinoma cells are the cells of 4T1 primary tumors that metastasized to the heart. Cells of 4THM tumors which metastasized to liver (4TLM) were previously isolated. Recently macroscopic brain metastasis in 4THM injected animals, were isolated to obtain a brain metastatic cell line (4TBM). Using an orthotopic mouse model differential characteristic of cells metastasized to heart (4THM), liver (4TLM), and brain (4TBM) were compared for ability to metastasize and expression of stem cell markers. We found that 4TLM cells produced significantly more lung and liver metastasis compared to 4TBM and 4THM cells. In vitro, proliferation as well as migration rate of 4TLM cells was also significantly higher than the other cell lines. Remarkably primary tumors formed by 4TLM cells expressed significant amounts of CD34, a marker for mesenchymal malignancies. Markers of epithelial-mesenchymal transition were expressed in all metastatic cells, but the degree of expression differed. Majorities of 4TLM, 4THM, and 4TBM cells were CD44+ CD24- whereas, 12 % of 4TLM cells also expressed membranous CD24. Conditioned mediums of non-metastatic 67NR breast tumors and cancer-associated fibroblasts inhibited growth of highly metastatic 4TLM cells. Malignant cells metastasized to brain were distinguished by membranous E-cadherin expression that was markedly higher in 4TBM cells grown as spheroids suggesting E-cadherin is required for brain metastasis. Differential features of heart, brain, and liver metastatic cells in a syngenic model was shown in this study for the first time. These findings not only provide a model to explore new treatment modalities, but also demonstrate differential features of cancer cells that originally homed to a certain organ, such as liver or brain.

Zinc (Zn) and adipose-derived mesenchymal stem cells (AD-MSCs) on MPTP-induced Parkinson's disease model: A comparative evaluation of behavioral and immunohistochemical results.

Parkinson's disease (PD) is characterized by the loss of dopaminergic neurons and sustained neuroinflammation due to microglial activation. Adipose tissue-derived mesenchymal stem cells (AD-MSCs) secrete neuroprotective factors to prevent neuronal damage. Furthermore, Zn regulates stem cell proliferation and differentiation and has immunomodulatory functions. Our in vivo study aimed to investigate whether Zn affects the activities of AD-MSCs in the MPTP-induced mouse model. Male C57BL/6 mice were randomly divided into six groups (n = 6): Control, Zn, PD, PD+Zn, PD+ (AD-MSC), PD+ (AD-MSC)+Zn. MPTP toxin (20 mg/kg) was dissolved in saline and intraperitoneally injected into experimental groups for two days with 12 h intervals. On the 3rd day, AD-MSCs were given to the right lateral ventricle of the PD+ (AD-MSC) and PD+ (AD-MSC)+Zn groups by stereotaxic surgery. Then, ZnSO4H2O was administered intraperitoneally for 4 days at 2 mg/kg. Seven days post MPTP injection, the motor activities of the mouse were evaluated. Then immunohistochemical analyzes were performed in SNpc. Our results showed that motor activity was lower in Group PD. AD-MSC and Zn administration have improved this impairment. MPTP caused a decrease in TH and BDNF expressions in dopaminergic neurons in Group PD. However, TH and BDNF expressions were more intense in the other groups. MCP-1, TGF-ß, and IL-10 expressions increased in administered groups compared to the Group PD. The present study indicates that Zn's individual and combined administration with AD-MSCs reduces neuronal damage in the MPTP-induced mouse model. In addition, anti-inflammatory responses that emerge with Zn and AD-MSCs may have a neuroprotective effect.

Olvanil activates sensory nerve fibers, increases T cell response and decreases metastasis of breast carcinoma.

BACKGROUND: Inactivation of sensory neurons expressing transient receptor potential vanilloid 1 (TRPV1) enhances breast cancer metastasis. Sensory neurons have profound effects on immune response to a wide range of diseases including cancer. Hence, activation of sensory nerves using feasible approaches such as specific TRPV1 agonists may inhibit breast cancer metastasis through neuroimmune pathways. TRPV1 agonists are considered for the treatment of pain and inflammatory diseases. METHODS: We here first determined the effects of four different TRPV1 agonists on proliferation of three different metastatic breast carcinoma cells since TRPV1 is also expressed in cancer cells. Based on the results obtained under in-vitro conditions, brain metastatic breast carcinoma cells (4TBM) implanted orthotopically into the mammary-pad of Balb-c mice followed by olvanil treatment (i.p.). Changes in tumor growth, metastasis and immune response to cancer cells were determined. RESULTS: Olvanil dose-dependently activated sensory nerve fibers and markedly suppressed lung and liver metastasis without altering the growth of primary tumors. Olvanil (5 mg/kg) systemically increased T cell count, enhanced intra-tumoral recruitment of CD8+ T cells and increased IFN-γ response to irradiated cancer cells and Con-A. Anti-inflammatory changes such as increased IL-10 and decrease IL-6 as well as S100A8+ cells were observed following olvanil treatment. CONCLUSIONS: Our results show that anti-metastatic effects of olvanil is mainly due to activation of neuro-immune pathways since olvanil dose used here is not high enough to directly activate immune cells. Furthermore, olvanil effectively depletes sensory neuropeptides; hence, olvanil is a good non-pungent alternative to capsaicin.

Effects of Melatonin and Doxorubicin on Primary Tumor And Metastasis in Breast Cancer Model.

BACKGROUND: Melatonin exerts oncostatic effects on breast cancer via immunomodulation and antioxidation. Doxorubicin is an effective chemotherapeutic agent, but parallel studies also provide ample evidence of an off-target effect of Doxorubicin in breast cancer patients. OBJECTIVE: Combinatorial use of doxorubicin and melatonin has not been comprehensively analyzed in breast cancer models. We hypothesized that the anti-oxidative, anti-proliferative and anti-inflammatory effects of melatonin could ameliorate the off-target effects of doxorubicin in breast cancer patients and enhance the anti-tumoral effects of doxorubicin. The goal of the study is to test this hypothesis in cancer cell lines and xenografted mice. METHODS: The effects of Melatonin and doxorubicin on the cell viability were evaluated in 4T1-Brain Metastatic Tumor (4TBM). Furthermore, the effects of melatonin and doxorubicin on the primary tumors and systemic metastasis were evaluated in the xenografted mice. Lung and liver tissues were removed and metastasis analyses were performed. The levels of p65, phospho-STAT3, CD11b+, GR1+, Ki67, and cleaved caspase-3 proteins were determined with immunohistochemistry and western blot analysis. We examined the effects of melatonin and Melatonin+Doxorubicin combination therapy on 4TBM cells. RESULTS: Our results showed that doxorubicin inhibited the proliferation of metastatic breast cancer cells while melatonin did not affect cells. Tumor growth and metastasis were markedly suppressed in melatonin alone and in combination with doxorubicin. The expression of CD11b+ and GR1+ proteins, which are indicators of myeloid-derived suppressor cells (MDSCs), were noted to be reduced in both primary tumor and metastatic tissues in melatonin and doxorubicin groups. CONCLUSION: The combination of melatonin with doxorubicin reduced primary tumor growth and distant metastasis. Based on these results, melatonin is a promising candidate for combinatory use with conventional chemotherapeutics for breast cancer treatment.

Role of melatonin in TLR4-mediated inflammatory pathway in the MTPT-induced mouse model.

Neuroinflammation has an essential role in various neurodegenerative diseases including Parkinson's disease (PD). Microglial activation as a result of neuroinflammation exacerbates the pathological consequences of the disease. The toxic effect of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) causes alpha-synuclein (α-synuclein) accumulation, which leads to dopaminergic neuron death in the MPTP-induced mouse model. Toll-like receptor 4 (TLR4) stimulates release of cytokine through NF-kB by activating glial cells, thus resulting in the death of dopaminergic neurons. Melatonin has the ability to cross the blood-brain barrier and protect neurons through anti-inflammatory properties. We hypothesized that melatonin could suppress TLR4-mediated neuroinflammation, decrease cytokine release due to the inflammatory response, and reduce dopaminergic neuron loss in the MPTP-induced mouse model. In the MPTP-induced mouse model, we aimed to assess the neuroinflammatory responses caused by TLR4 activation as well as the effect of melatonin on these responses. Three-month-old male C57BL/6 mice were randomly divided into five groups; Control (Group-C), Sham (Group-S), Melatonin-treated (Group-M), MPTP-injected (Group-P), and MPTP + melatonin-injected (Group-P + M). MPTP toxin (20 mg/kg) was dissolved in saline and intraperitoneally (i.p.) injected to mice for two days with 12 h intervals. The total dose per mouse was 80 mg/kg. Melatonin was administered (20 mg/kg) intraperitoneally to Group-M and Group-P + M twice a day for five days. Eight days after starting the experiment, the motor activities of mice were evaluated by locomotor activity tests. The effects on dopamine neurons in the SNPc was determined by tyrosine hydroxylase (TH) immunohistochemistry. TLR4, α-synuclein, and p65 expression was evaluated by immunostaining as well. The amount of TNF-alpha in the total brain was evaluated by western blot analysis. In our results seen that locomotor activity was lower in Group-P compared to Group-C. However, melatonin administration was improved this impairment. MPTPcaused decrease in TH immuno-expression in dopaminergic neurons in Group-P. TLR4 (p < 0.001), α-synuclein (p < 0.001), and p65 (p < 0.01) immuno-expressions were also decreased in Group-P+M compared to Group-P (using MPTP). TNF-α expression was lower in Group-C, Group-S, Group-M, and Group-P+M, when compared to Group-P (p < 0.0001) due to the absence of inflammatory response. In conclusion, our study revealed that melatonin administration reduced α-synuclein aggregation and TLR4-mediated inflammatory response in the MPTP-induced mouse model.

Comparison of the therapeutic effects of erythropoietin and acetyl-l-carnitine on sciatic nerve injury in rats.

AIM: We aimed to investigate the effects of erythropoietin, acetyl-l-carnitine, and their combination on nerve regeneration in experimental peripheral nerve injury. METHODS: Rats were randomly divided into five groups - sham-operated (S), sciatic nerve crush injury (C), C + acetyl-l-carnitine (ALCAR), C + erythropoietin (EPO), and C + EPO + ALCAR. ALCAR (50 mg/kg/day) was administered intraperitoneally, and EPO (5000 U/kg) was injected subcutaneously for 10 days. Functional recovery was evaluated using walking track analysis (sciatic functional index [SFI]), somatosensory evoked potentials (SEPs), thiobarbituric acid reactive substance (TBARS) assay, and caspase-3 and S100 immunoreactivities. RESULTS: In SFI analyses, delayed functional recovery was observed in the C group, whereas the functional recovery of rats treated with EPO and ALCAR significantly improved. The latencies of the SEP components were significantly prolonged in C group. In the treatment groups (C + EPO, C + ALCAR, and C + EPO + ALCAR), all recorded values of SEP components significantly decreased. TBARS levels in C group were significantly higher than those in the S group. EPO and ALCAR administration significantly decreased TBARS levels. Caspase-3 immunoreactivity was increased in the C group, whereas it was decreased in the treatment groups. S100 immunolabelling was significantly decreased in the C group. EPO and ALCAR administration caused an increase in the amount of S100-positive cells in all treatment groups. CONCLUSION: EPO and ALCAR administration could accelerate sciatic nerve repair by reducing apoptosis and lipid peroxidation and promoting myelinization. Although both EPO and ALCAR had positive effects on nerve healing, their combined efficacy had no statistically significant effect on peripheral nerve regeneration.