Microfluidic-based technologies in cancer liquid biopsy: Unveiling the role of horizontal gene transfer (HGT) materials.

Liquid biopsy includes the isolating and analysis of non-solid biological samples enables us to find new ways for molecular profiling, prognostic assessment, and better therapeutic decision-making in cancer patients. Despite the conventional theory of tumor development, a non-vertical transmission of DNA has been reported among cancer cells and between cancer and normal cells. The phenomenon referred to as horizontal gene transfer (HGT) has the ability to amplify the advancement of tumors by disseminating genes that encode molecules conferring benefits to the survival or metastasis of cancer cells. Currently, common liquid biopsy approaches include the analysis of extracellular vesicles (EVs) and tumor-free DNA (tfDNA) derived from primary tumors and their metastatic sites, which are well-known HGT mediators in cancer cells. Current technological and molecular advances expedited the high-throughput and high-sensitive HGT materials analyses by using new technologies, such as microfluidics in liquid biopsies. This review delves into the convergence of microfluidic-based technologies and the investigation of Horizontal Gene Transfer (HGT) materials in cancer liquid biopsy. The integration of microfluidics offers unprecedented advantages such as high sensitivity, rapid analysis, and the ability to analyze rare cell populations. These attributes are instrumental in detecting and characterizing CTCs, circulating nucleic acids, and EVs, which are carriers of genetic cargo that could potentially undergo HGT. The phenomenon of HGT in cancer has raised intriguing questions about its role in driving genomic diversity and acquired drug resistance. By leveraging microfluidic platforms, researchers have been able to capture and analyze individual cells or genetic material with enhanced precision, shedding light on the potential transfer of genetic material between cancer cells and surrounding stromal cells. Furthermore, the application of microfluidics in single-cell sequencing has enabled the elucidation of the genetic changes associated with HGT events, providing insights into the evolution of tumor genomes. This review also discusses the challenges and opportunities in studying HGT materials using microfluidic-based technologies. In conclusion, microfluidic-based technologies have significantly advanced the field of cancer liquid biopsy, enabling the sensitive and accurate detection of HGT materials. As the understanding of HGT's role in tumor evolution and therapy resistance continues to evolve, the synergistic integration of microfluidics and HGT research promises to provide valuable insights into cancer biology, with potential implications for precision oncology and therapeutic strategies.

Carbon dot incorporated mesoporous silica nanoparticles for targeted cancer therapy and fluorescence imaging.

A new and efficient theranostic nanoplatform was developed via a green approach for targeted cancer therapy and fluorescence imaging, without the use of any anticancer chemotherapeutic drugs. Toward this aim, monodisperse and spherical mesoporous silica nanoparticles (MSNs) of approximately 50 nm diameter were first synthesized using the sol-gel method and loaded with hydrothermally synthesized anticancer carbon dots (CDs). The resulting MSNs-CDs were then functionalized with chitosan and targeted by an anti-MUC1 aptamer, using the glutaraldehyde cross-linker, and fully characterized by TEM, FE-SEM, EDS, FTIR, TGA, XRD, and BET analysis. Potent and selective anticancer activity was obtained against MCF-7 and MDA-MB-231 cancer cells with the maximum cell mortalities of 66.2 ± 1.97 and 71.8 ± 3%, respectively, after 48 h exposure with 100 µg mL-1 of the functionalized MSNs-CDs. The maximum mortality of 40.66 ± 1.3% of normal HUVEC cells was obtained under the same conditions. Based on the results of flowcytometry analysis, the apoptotic mediated cell death was recognized as the main anticancer mechanism of the MSNs-CDs. The fluorescence imaging of MCF-7 cancer cells was also studied after exposure with MSNs-CDs. The overall results indicated the high potential of the developed nanoplatform for targeted cancer theranostics.

Facile, rapid and efficient isolation of circulating tumor cells using aptamer-targeted magnetic nanoparticles integrated with a microfluidic device.

Facile and sensitive detection and isolation of circulating tumor cells (CTCs) was achieved using the aptamer-targeted magnetic nanoparticles (Apt-MNPs) in conjugation with a microfluidic device. Apt-MNPs were developed by the covalent attachment of anti-MUC1 aptamer to the silica-coated magnetic nanoparticles via the glutaraldehyde linkers. Apt-MNPs displayed high stability and functionality after 6 months of storage at 4 °C. The specific microfluidic device consisting of mixing, sorting and separation modules was fabricated through conventional photo- and soft-lithography by using polydimethylsiloxane. The capture efficiency of Apt-MNPs was first studied in vitro on MCF-7 and MDA-MB-231 cancer cell lines in the bulk and microfluidic platforms. The cell capture yields of more than 91% were obtained at the optimum condition after 60 minutes of exposure to 50 µg mL-1 Apt-MNPs with 10 to 106 cancer cells in different media. CTCs were also isolated efficiently from the blood samples of breast cancer patients and successfully propagated in vitro. The isolated CTCs were further characterized using immunofluorescence staining. The overall results indicated the high potential of the present method for the detection and capture of CTCs.

Three-Dimensional in Vitro Models: A Promising Tool To Scale-Up Breast Cancer Research.

Common models used in breast cancer studies, including two-dimensional (2D) cultures and animal models, do not precisely model all aspects of breast tumors. These models do not well simulate the cell-cell and cell-stromal interactions required for normal tumor growth in the body and lake tumor like microenvironment. Three-dimensional (3D) cell culture models are novel approaches to studying breast cancer. They do not have the restrictions of these conventional models and are able to recapitulate the structural architecture, complexity, and specific function of breast tumors and provide similar in vivo responses to therapeutic regimens. These models can be a link between former traditional 2D culture and in vivo models and are necessary for further studies in cancer. This review attempts to summarize the most common 3D in vitro models used in breast cancer studies, including scaffold-free (spheroid and organoid), scaffold-based, and chip-based models, particularly focused on the basic and translational application of these 3D models in drug screening and the tumor microenvironment in breast cancer.

Effect of Anti-Podoplanin on Malignant Glioma Cell Viability, Invasion and Tumor Cell-Induced Platelet Aggregation.

BACKGROUND: The transmembrane receptor podoplanin (PDPN) is a platelet aggregation-inducing factor, which is widely expressed in various malignant tumors such as squamous cell carcinomas, mesotheliomas, glioblastomas. Podoplanin regulates a pathway leading to cell invasion and migration. Glioblastoma multiforme (GBM) is the most aggressive and invasive tumor of the central nervous system. A high level of PDPN expression has been reported to be associated with reduced survival, cancer aggression and migration. Aim of study to determine the effect of anti-podoplanin antibody on cell-platelet aggregation, cell invasion and viability in Uppsala 87 malignant glioma (U87MG) cell lines. METHODS: The expression of podoplanin on U87MG cells was measured using flowcytometry. The dimethyl-thiazol diphenyl-tetrazolium bromide (MTT) assay was used to measure U87MG cell proliferation after treatment by anti-podoplanin monoclonal antibody (NZ-1.3). The invasion of cancer cells was assessed by using a novel microfluidic based assay. RESULTS: The results show reduction of cell viability and cell migration after treatment with anti-podoplanin antibody. After co- treatment of U87MG cells and platelets with and without anti-podoplanin antibody, the cell-platelet aggregation was significantly reduced in anti-podoplanin treated cell. CONCLUSIONS: Podoplanin is involved in aggregation of gliobastoma cells, and their viability and invasion and its neutralizing antibody can inhibit this process. So, blocking of podoplanin may be representing a promising therapeutic approach to glioblastoma multiform cancer therapy.

Effects of Methadone on the Toll-like Receptor 4 Expression in Human Non-Small Cell Lung Carcinoma A549 Cell Line Using In-silico and In vitro Techniques.

Background: In this study, the effects of methadone and naloxone on the expression of toll-like receptor 4 (TLR4) gene have been evaluated in human non-small cell lung carcinoma A549 cell line migration using in-silico and in vitro techniques. Materials and Methods: Lung cancer A549 cell cultures were stimulated for 24 h with methadone (5, 10, and 20 µM) and naloxone (20 and 40 µM) concentrations. The level of TLR4 expression was determined by the quantitative real-time polymerase chain reaction. Migration of the A549 cells was investigated after a 4-h incubation period with methadone using the Boyden Chamber assay. Results: Migration rate of the A549 cells treated with 5 (P < 0.05) and 20 (P < 0.01) µM methadone was, respectively, increased and decreased with 20 µM naloxone (P < 0.05). Furthermore, the TLR4 expression was enhanced with 5 (P < 0.05) and 20 (P < 0.01) µM methadone and decreased with 20 (P < 0.05) and 40 µM naloxone (P < 0.01). In addition, in silico docking analysis revealed docking of methadone to MD-2 and TLR4. Conclusion: According to the present DATA, methadone affects the TLR4 expression. It may however cause adverse consequences by increasing the TLR4 expression. Therefore, the useful analgesic properties of methadone should be separated from the unwanted TLR4-mediated side effects.

A review on the role of CAFs and CAF-derived exosomes in progression and metastasis of digestive system cancers.

Cancers evolve as a result of the accelerated proliferation of cancer cells in a complicated, enriched, and active microenvironment. Tumor microenvironment (TME) components are the master regulators of any step of cancer development. The tumor microenvironment is composed of many cellular and noncellular components that contribute to the evolution of cancer cells. Cancer-associated fibroblasts (CAFs) are activated fibroblasts in the TME that implicate in tumor progression and metastasis dissemination through secretion of oncogenic factors which are carried to the secondary metastatic sites through exosomes. In this review, we aimed to assess the role of CAF-derived exosomes in TME construction and pre-metastatic niche formation in different cancers of the digestive system in order to better understand some important mechanisms of metastasis and provide possible targets for clinical intervention. This review article is divided into two thematic parts explaining the general mechanisms of pre-metastatic niche formation and metastasis and the role of CAF-derived exosomes in different digestive system cancers including colorectal, gastric, esophageal, pancreatic, and liver cancers.

AC electrokinetic isolation and detection of extracellular vesicles from dental pulp stem cells: Theoretical simulation incorporating fluid mechanics.

Extracellular vesicles (EVs) are cell-derived nanoscale vesicles involved in intracellular communication and the transportation of biomarkers. EVs released by mesenchymal stem cells have been recently reported to play a role in cell-free therapy of many diseases. However, the demand for better research tools to replace the tedious conventional methods used to study EVs is getting stronger. EVs' manipulation using alternating current (AC) electrokinetic forces in a microfluidic device has appeared to be a reliable and sensitive diagnosis and trapping technique. Given that different AC electrokinetic forces may contribute to the overall motion of particles and fluids in a microfluidic device, EVs' electrokinetic trapping must be examined considering all dominant forces involved depending on the experimental conditions. In this paper, AC electrokinetic trapping of EVs using an interdigitated electrode arrays is investigated. A 2D numerical simulation incorporating the two significant AC electrokinetic phenomena (Dielectrophoresis and AC electroosmosis) has been performed. Theoretical predictions are then compared with experimental results and allow for a plausible explanation of observations inconsistent with DEP theory. It is demonstrated that the inconsistencies can be attributed to a significant extent to the contribution of the AC electroosmotic effect.

A simple, low cost and reusable microfluidic gradient strategy and its application in modeling cancer invasion.

Microfluidic chemical gradient generators enable precise spatiotemporal control of chemotactic signals to study cellular behavior with high resolution and reliability. However, time and cost consuming preparation steps for cell adhesion in microchannels as well as requirement of pumping facilities usually complicate the application of the microfluidic assays. Here, we introduce a simple strategy for preparation of a reusable and stand-alone microfluidic gradient generator to study cellular behavior. Polydimethylsiloxane (PDMS) is directly mounted on the commercial polystyrene-based cell culture surfaces by manipulating the PDMS curing time to optimize bonding strength. The stand-alone strategy not only offers pumpless application of this microfluidic device but also ensures minimal fluidic pressure and consequently a leakage-free system. Elimination of any surface treatment or coating significantly facilitates the preparation of the microfluidic assay and offers a detachable PDMS microchip which can be reused following to a simple cleaning and sterilization step. The chemotactic signal in our microchip is further characterized using numerical and experimental evaluations and it is demonstrated that the device can generate both linear and polynomial signals. Finally, the feasibility of the strategy in deciphering cellular behavior is demonstrated by exploring cancer cell migration and invasion in response to chemical stimuli. The introduced strategy can significantly decrease the complexity of the microfluidic chemotaxis assays and increase their throughput for various cellular and molecular studies.

The dysbiosis signature of Fusobacterium nucleatum in colorectal cancer-cause or consequences? A systematic review.

Colorectal cancer (CRC) is the third most common cause of cancer globally and the fourth attributable cause of mortality and morbidity due to cancer. An emerging factor contributing to CRC is the gut microbiota and the cellular changes associated with it. Further insights on this may help in the prevention, diagnosis and new therapeutic approaches to colorectal cancer. In most cases of CRC, genetic factors appear to contribute less to its aetiology than environmental and epigenetic factors; therefore, it may be important to investigate these environmental factors, their effects, and the mechanisms that may contribute to this cancer. The gut microbiota has recently been highlighted as a potential risk factor that may affect the structural components of the tumor microenvironment, as well as free radical and enzymatic metabolites directly, or indirectly. Many studies have reported changes in the gut microbiota of patients with colorectal cancer. What is controversial is whether the cancer is the cause or consequence of the change in the microbiota. There is strong evidence supporting both possibilities. The presence of Fusobacterium nucleatum in human colorectal specimens has been demonstrated by RNA-sequencing. F. nucleatum has been shown to express high levels of virulence factors such as FadA, Fap2 and MORN2 proteins. Our review of the published data suggest that F. nucleatum may be a prognostic biomarker of CRC risk, and hence raises the potential of antibiotic treatment of F. nucleatum for the prevention of CRC.

Therapeutic inhibition of microRNA-21 (miR-21) using locked-nucleic acid (LNA)-anti-miR and its effects on the biological behaviors of melanoma cancer cells in preclinical studies.

BACKGROUND: Melanoma is a cancer that has a high mortality rate in the absence of targeted therapy. Conventional therapies such as surgery, chemotherapy, and radiotherapy are associated with poor prognosis. The expression of miR-21 appears to be of clinical importance, and the regulation of its expression appears to be an opportunity for treatment. METHODS: In this current study, we aimed to evaluate the effects of miR-21 inhibition in- vitro and in-vivo. In-vitro studies have investigated LNA-anti-miR-21 in mouse melanoma cells (B16F10), and in-vivo studies have proposed a model of melanoma in male C57BL/6 mice. To evaluate the anticancer effects of LNA-anti-miR-21, a QRT-PCR analysis was performed using the 2-ΔΔCT method to determine the degree of inhibition of oncomiR-21. The MTT test, propidium iodide/AnnexinV in-vitro, and tumor volume measurement using the QRT-PCR test with the 2-ΔΔCT method were used to estimate the inhibition of miR-21 and the expression of downstream genes including: SNAI1, Nestin (Nes), Oct-4, and NF-kB following miR-21 inhibition. Finally, immunohistochemistry was conducted for an in-vivo animal study. RESULTS: MiR-21 expression was inhibited by 80% after 24 h of B16F10 cell line transfection with LNA-anti-miR-21. The MTT test showed a significant reduction in the number of transfected cells with LNA-anti-miR-21. The transfected cells showed a significant increase in apoptosis in comparison with the control and scrambled LNA groups. According to our in vivo findings, anti-miR-21 could reduce tumor growth and volume in mice receiving intraperitoneal anti-miR after 9 days. The expression of the SNAI1gene was significantly reduced compared to the controls. Immunohistochemical analysis showed no change in CD133 and NF-kB markers. CONCLUSION: Our findings suggest LNA-anti-miR-21 can be potentially used as an anticancer agent for the treatment of melanoma.

Evaluation of rs1982073 polymorphism of transforming growth factor-ß1 in glioblastoma.

BACKGROUND: Glioblastoma (GBM) is the most common and invasive form of primary malignant brain tumors, with a survival rate of about 1 year. Transforming growth factor-ß1 (TGF-ß1) plays a very important role in tissue homeostasis and cancers. It seems that polymorphism of T29C (L10P, rs1982073, or rs1800470), which has been studied in various cancers such as breast and colon, creates the significant differences plays an important role in GBM prognosis and treatment. In this study, we evaluated the effect of T29C (rs1982073) polymorphism of TGF-ß1 gene in GBM. MATERIALS AND METHODS: This study was conducted on 100 cases of GBM including 47 paraffin-embedded brain tissue samples and 53 blood samples from another 53 GBM patients, who was under therapy, and 150 were controls. The TGF-ß rs1982073 single-nucleotide polymorphism (SNP) was identified by the NCBI and genotyping was performed by high-resolution melt (HRM) assay. Melt curves from HRM which suspected to SNP were selected and subjected to direct sequencing. Finally, the collected data were entered into the SPSS software (Version. 20) and mean ± standard deviation or n (%) was used to show the data. RESULTS: The mean age in GBM group was 51.63 ± 13.27 years. Accordingly, the two groups were matched in terms of age and gender (P > 0.05). The frequency of GG genotype was significantly higher in GBM patients. In contrast, although the frequency of AG genotype was higher in GBM group, it was not statistically significant. Furthermore, the presence of G allele was significantly more frequent than A allele in GBM patients. CONCLUSION: Findings of the present study supports that the Pro10Leu, rs1982073, or rs1800470 SNP in TGF-ß1 is found to be expressed significantly more in GBM patients as it was found in breast cancer.

Maprotiline inhibits COX2 and iNOS gene expression in lipopolysaccharide-stimulated U937 macrophages and carrageenan-induced paw edema in rats.

Maprotiline, a tetracyclic antidepressant, is used for the management of mental disorders and various types of chronic pain. In our previous work, we found the inhibitory effect of maprotiline on inflammatory mediator's expression like tumor necrosis factor α (TNF-α and interleukin 1ß (IL-1ß. As part of that study, we sought to evaluate the effect of maprotiline on the expression of some inflammatory mediators such as cyclooxygenases 2 (COX2) and inducible nitric oxide synthase (iNOS). For this reason we used an in vitro model system of lipopolysaccharide (LPS)-stimulated human U937 macrophages and also an in vivo model of carrageenan-induced paw edema in rats. We measured the expression of these genes by quantitative RT-real time PCR. The expression of COX2 and iNOS significantly decreased by maprotiline in U937 macrophages and carrageenan-induced paw inflammation in rats. Our finding also confirmed that intraperitoneal (i.p.) injection of maprotiline inhibited carrageenan-induced paw edema. Moreover, maprotiline significantly decreased the migration of polymorphonuclear (PMN) leukocytes to the site of inflammation. The results of the present study provide further evidence for the anti-inflammatory effect of maprotiline. This effect appears to be mediated by down regulation of inflammatory genes. Further studies are needed to evaluate the complex cellular and molecular mechanisms of maprotiline.

Evaluation of the Effect of Antidepressant Drug, Fluvoxamine, on Cyclooxygenase-2 Protein Expression in Lipopolysaccharide-stimulated Macrophages.

BACKGROUND: Fluvoxamine, a well-known selective serotonin reuptake inhibitor, is used for the management of mental disorders and various types of chronic pain. In our previous study, we found the inhibitory effect of fluvoxamine on inflammatory mediator's expression. In the line of the indicated study, we sought to evaluate the effect of fluvoxamine on the expression of some inflammatory mediators such as cyclooxygenase-2 (COX-2). MATERIALS AND METHODS: An in vitro model system of lipopolysaccharide-stimulated human U937 macrophages was used. The expression of COX-2 protein was measured by flow cytometry. RESULTS: The expression of COX-2 significantly decreased by fluvoxamine in U937 macrophages. CONCLUSION: The results of the present study provide further evidence for the anti-inflammatory effect of fluvoxamine. This effect appears to be mediated by the downregulation of inflammatory genes. Further studies are needed to evaluate the complex cellular and molecular mechanisms of fluvoxamine.

Evaluation of rs1957106 Polymorphism of NF-κBI in Glioblastoma Multiforme in Isfahan, Iran.

BACKGROUND: The kB family of nuclear factor (NF-κB) is a series of transcription factors that plays a key role in regulation of immunity, cell growth, and apoptosis and is considered as the main downstream component of epidermal growth factor receptor for which there are evidence of excessive activity in most cases of glioblastoma multiform (GBM). Thus, the current information has gained evidence on NF-κBIA tumor suppressor role in GMB. SNP rs1957106 was diagnosed as a new polymorphism which affected the expression of NF-κBI and causes activation of NF-κB in GBM patients. MATERIALS AND METHODS: This study was conducted on 100 cases of GBM including 47 paraffin-embedded brain tissue samples and 53 blood samples from another 53 GBM patients and 150 controls. The NF-κBI rs1957106 SNP was identified by the NCBI, and genotyping was performed by high-resolution melt (HRM) assay. Melt curves from HRM which suspected to single-nucleotide polymorphism (SNP) were selected and subjected to direct sequencing. RESULTS: The distribution of allele A of NF-κß gene in patients with GBM with 31% was not significantly different from healthy participants (27.3%) (P = 0.375). Furthermore, the distribution of AG and GG genotypes in comparison with AA genotypes did not show a significant correlation with GBM incidence (P > 0.05). CONCLUSION: Findings of the present study provide evidence that the rs1957106 SNP in NF-κBIA is found more in GBM patients, but it was not statistically significant. As there are conflicting studies showing significant higher rate of this SNP in GBM, further study is suggested.

Association between autocrine motility factor receptor gene polymorphism (rs2440472, rs373191257) and glioblastoma multiform in a representative Iranian population.

BACKGROUND: Glioblastoma multiform (GBM) is the most common and most malignant of the glial tumors that begins primarily in brain tissue. Genetic background could be considered as an important predisposing factor in GBM. Autocrine motility factor receptor (AMFR) is a cytokine receptor that participates in a lot of physiologic and pathologic processes like: Cellular motility and metastasis. So, it seems that this protein has an essential role in pathophysiology of several cancers and could be a potential diagnostic and or therapeutic target in GBM. The aim of this study is to investigate the association of AMFR (rs2440472, rs373191257) gene polymorphism and GBM in a representative Iranian population. MATERIALS AND METHODS: This study includes 81 cases of GBM and 117 control subjects. After DNA extraction, polymerase chain reaction - high resolution melting reaction was performed. For each single nucleotide polymorphisms, 12 samples were selected for sequencing. Data was analyzed using Chi-square test and Logistic regression. RESULTS: For rs2440472, frequency of GG genotype in the case group was increased compared to the control group (51.9% vs. 34.2% respectively, P = 0.013). After adjusting for sex and age by logistic regression our results were the same (P = 0.017, odds ratio = 2.056). Allelic frequencies for rs2440472 among cases and controls were not significantly different (P = 0.058). For rs373191257, genotypic and allelic frequencies were not significantly different between two groups. CONCLUSION: Our results showed the possible association between the AMFR rs2440472 gene polymorphism with susceptibility to GBM.

The Study of Association Between Polymorphism of TNF-α Gene's Promoter Region and Recurrent Pregnancy Loss.

BACKGROUND: According to the literature review, polymorphisms of tumor necrosis factor alpha's (TNF-α) promoter region are probably the genetic risk factors of recurrent pregnancy loss. This study has investigated five single nucleotide polymorphisms in the TNF-α gene's promoter region to evaluate their relationship with recurrent pregnancy loss disorder. METHODS: Blood samples were taken from 65 women with recurrent pregnancy loss (Case group) and 65 healthy women with a history of successful pregnancy (Control group). Polymerase chain reaction with high resolution melting (PCR-HRM) analysis was done to determine the promoter region of -308G/A, -850T/C, -238G/A, -1031T/C and -863A/C TNF-α polymorphisms. The data were assessed using logistic regression models. P values less than 0.05 were considered statistically significant. RESULTS: Significant associations were found between recurrent pregnancy loss and -863C/A (p=0.000), -308G/A (p=0.045), and -238G/A (p=0.034) polymorphisms. TNF-α polymorphisms of -863C and -238G may be susceptible factors of recurrent pregnancy loss cases. The -308G polymorphism has an important role in maintaining pregnancy. CONCLUSION: The -863C/A and -238G/A TNF-α polymorphisms are possible genetic risk factors of recurrent pregnancy loss and might be its predictive markers.

Non-viral Delivery Systems for Breast Cancer Gene Therapy.

INTRODUCTION: The ever-evolving field of gene therapy promises several innovative treatments for cancer. Advances in genetic modification of tumor cells and micro environment have led to the development of more effective therapeutic strategies with fewer side effects. MATERIALS & METHODS: The development of effective delivery system challenges, remains. Non-viral vectors are interesting due to their bio-safety and their ability to transfer different types of nucleic acids. Examples of these techniques are the use of oligonucleoides, liposomes, nanoparticles, inorganic material, and engineered stem cells. CONCLUSION: In this review, we focus on recent advances in the intracellular delivery of DNA and siRNA to the cancer cells with emphasis on breast cancer.

Effects of doxepin on gene expressions of Bcl-2 family, TNF-α, MAP kinase 14, and Akt1 in the hippocampus of rats exposed to stress.

Stress is one of the effective factors in the development of depressive disorders that performs some parts of its effects by affecting hippocampus. Since doxepin has been shown to have neuroprotective effects, in this study, we focused on the effects of doxepin on the expression of involved genes in neuronal survival and plasticity in the rat hippocampus following chronic stress. Male Wistar rats were divided into four groups, the control, the stress, the stress-doxepin 1 mg/kg and the stress-doxepin 5 mg/kg, respectively. To induce stress, the rats were placed within adjustable restraint chambers for 6 h/day, for 21 days. Before daily induction of the stress, rats received an i.p. injection of doxepin. At the end of experiments, expression of Bax, Bad, Bcl-2, tumor necrosis factor alpha (TNF-α), mitogen-activated protein kinase 14 (MAPK14) and serine-threonine protein kinase AKT1 genes were detected by reverse transcription polymerase chain reaction (RT-PCR) in the hippocampus. Results showed significant enhancements in expression of Bax, Bad and Bcl-2 genes in the stressed rats, whereas expression of TNF-α, MAPK14, and AKT1 genes didn't show significant differences. Doxepin could decrease the expression of Bax and Bad genes in the stress group, but had no significant effects on the expression of other genes. The present findings indicated that doxepin can probably change the pattern of gene expression in the hippocampus to maintain neurons against destructive effects of stress.