Dynamical changes in the expression of GABAergic and purinergic components occur during the polarization of THP-1 monocytes to proinflammatory macrophages.

The monocytes are key components of innate immunity, as they can differentiate into phagocytic cells or macrophages with proinflammatory or anti-inflammatory phenotypes. The gamma-aminobutyric acid (GABA) and adenosine triphosphate (ATP), two known neurotransmitters, are two environmental signals that contribute to the differentiation of monocytes into macrophages and their subsequent polarization into proinflammatory M1 and anti-inflammatory M2 macrophages. Although monocytes and macrophages express proteins related to GABA and ATP-mediated response (GABAergic and purinergic systems, respectively), it is unknown whether changes in their expression occur during monocyte activation or their differentiation and polarization into macrophages. Therefore, we evaluated the expression levels of GABAergic and purinergic signaling components in the THP-1 monocyte cell line and their changes during monocyte activation, differentiation, and polarization to M1 proinflammatory macrophages. Our results showed that activated monocytes are characterized by increased expression of two GABAergic components, the GABA transporter 2 (GAT-2) and the glutamic acid decarboxylase (GAD)-67, an enzyme involved in GABA synthesis. Also, monocytes showed a pronounced expression of the purinergic receptors P2X4 and P2X7. Interestingly, during differentiation, monocytes increased the expression of the ß2 subunit of GABA A-type receptor (GABA-AR), while the purinergic receptors P2X1 and P2X1del were reduced. In contrast, proinflammatory M1 macrophages showed a reduced expression in the α4 subunit of GABA-AR and GAD67, while P2X4 and P2X7 were overexpressed. These results indicate that dynamical changes in the GABAergic and purinergic components occur during the transition from monocytes to macrophages. Since GABA and ATP are two neurotransmitters, our results suggest that monocytes and macrophages respond to neurotransmitter-induced stimulation and may represent a path of interaction between the nervous and immune systems during peripheral inflammation and neuroinflammation development.

Revealing the contribution of astrocytes to glutamatergic neuronal transmission.

Research on glutamatergic neurotransmission has focused mainly on the function of presynaptic and postsynaptic neurons, leaving astrocytes with a secondary role only to ensure successful neurotransmission. However, recent evidence indicates that astrocytes contribute actively and even regulate neuronal transmission at different levels. This review establishes a framework by comparing glutamatergic components between neurons and astrocytes to examine how astrocytes modulate or otherwise influence neuronal transmission. We have included the most recent findings about the role of astrocytes in neurotransmission, allowing us to understand the complex network of neuron-astrocyte interactions. However, despite the knowledge of synaptic modulation by astrocytes, their contribution to specific physiological and pathological conditions remains to be elucidated. A full understanding of the astrocyte's role in neuronal processing could open fruitful new frontiers in the development of therapeutic applications.

Clinical utility of ABCB1 and ABCG2 genotyping for assessing the clinical and pathological response to FAC therapy in Mexican breast cancer patients.

PURPOSE: Resistance to neoadjuvant chemotherapy with 5-fluorouracil, doxorubicin, and cyclophosphamide (FAC) in some patients with locally advanced breast cancer remains one of the main obstacles to first-line treatment. We investigated clinical and pathological responses to FAC neoadjuvant chemotherapy in Mexican women with breast cancer and their possible association with SNPs present in ABC transporters as predictors of chemoresistance. MATERIALS: A total of 102 patients undergoing FAC neoadjuvant chemotherapy were included in the study. SNP analysis was performed by RT-PCR from genomic DNA. Two SNPs were analyzed: ABCB1 rs1045642 (3435 C > T) and ABCG2 rs2231142 (421 G > T). RESULTS: In clinical response evaluation, significant associations were found between the ABCB1 C3435T genotype and breast cancer chemoresistant and chemosensitive patients (p < 0.05). In the early clinical response, patients with genotype C/C or C/T were more likely to be chemosensitive to neoadjuvant therapy than patients with genotype T/T (OR = 4.055; p = 0.0064). Association analysis between the ABCB1 gene polymorphism and the pathologic response to FAC chemotherapy showed that the C/C + C/T genotype was a protective factor against chemoresistance (OR = 3.714; p = 0.0104). Polymorphisms in ABCG2 indicated a lack of association with resistance to chemotherapy (p = 0.2586) evaluating the clinical or pathological response rate to FAC neoadjuvant chemotherapy. CONCLUSION: The early clinical response and its association with SNPs in the ABCB1 transporter are preserved until the pathological response to neoadjuvant chemotherapy; therefore, it could be used as a predictor of chemoresistance in locally advanced breast cancer patients of the Mexican population.

Chemoresistance in Breast Cancer Patients Associated With Changes in P2X7 and A2A Purinergic Receptors in CD8+ T Lymphocytes.

Breast cancer (BRCA) is the most frequent cancer type that afflicts women. Unfortunately, despite all the current therapeutic strategies, many patients develop chemoresistance hampering the efficacy of treatment. Hence, an early indicator of therapy efficacy might aid in the search for better treatment and patient survival. Although emerging evidence indicates a key role of the purinergic receptors P2X7 and A2A in cancer, less is known about their involvement in BRCA chemoresistance. In this sense, as the chemotherapeutic treatment stimulates immune system response, we evaluated the expression and function of P2X7 and A2A receptors in CD8+ T cells before and four months after BRCA patients received neoadjuvant chemotherapy. The results showed an increase in the levels of expression of P2X7 and a decrease in the expression of A2A in CD8+ T cells in non-chemoresistant (N-CHR) patients, compared to chemoresistant (CHR) patients. Interestingly, in CHR patients, reduced expression of P2X7 occurs along with a decrease in the CD62L shedding and the production of IFN-γ. In the case of the A2A function, the inhibition of IFN-γ production was not observed after chemotherapy in CHR patients. A possible relationship between the modulation of the expression and function of the P2X7 and A2A receptors was found, according to the molecular subtypes, where the patients that were triple-negative and human epidermal growth factor receptor 2 (HER2)-enriched presented more alterations. Comorbidities such as overweight/obesity and type 2 diabetes mellitus (T2DM) participate in the abnormalities detected. Our results demonstrate the importance of purinergic signaling in CD8+ T cells during chemoresistance, and it could be considered to implement personalized therapeutic strategies.

P2X4 receptor as a modulator in the function of P2X receptor in CD4+ T cells from peripheral blood and adipose tissue.

Obesity is characterized by immune cell infiltration and inflammation. Purinergic receptors such as P2X1, 4 and 7 are expressed on immune cells and their activation contributes with an inflammatory response. However, the simultaneous expression of P2X1, 4 and 7 during overweight or obesity have not been described. Therefore, the aim of this study was to determine single and simultaneously expression and function of the P2X1, 4 and 7 receptors in lymphocytes and CD4 + T cells from peripheral blood (PB) and adipose tissue (AT). Our results showed a higher expression of the P2X4 receptor on CD4 + T cells from PB regarding P2X7 and P2X1 receptor expression. In addition, P2X4 receptor expression on CD4 + T cells from PB and AT was increased in individuals with BMI ≥ 25 Kg/m2. Moreover, a higher simultaneous expression of the P2X4 and P2X7 receptors on CD4 + T cells from AT compared to CD4 + T cells expressing P2X1 and P2X7 receptors simultaneously. Besides, CD4 + T cells expressing P2X4 and P2X7 receptors from PB and AT were augmented in individuals with BMI ≥ 25 Kg/m2. In addition, the percentage of lymphocytes and also CD4 + T cells expressing P2X4 receptor were elevated both in PB and AT compared to cells expressing P2X7 or P2X1. However, CD4 + T cells expressing P2X4 and P2X7 were augmented in AT compared to PB. The function of the receptors showed a lower shedding of CD62 L in adipose tissue mononuclear cells (ATMC) compared with peripheral blood mononuclear cells (PBMC) and a greater participation of P2X4 in the mobilization of intracellular calcium. We concluded that it was possible to determine for the first time the simultaneous expression of purinergic receptors in ATMC, where the P2X4 receptor has a greater participation in the activation of CD4 + T cells possibly modulating the function of the other two receptors.

Diminished levels of regulatory T cell subsets (CD8+Foxp3, CD4+Foxp3 and CD4+CD39+Foxp3) but increased Foxp3 expression in adipose tissue from overweight subjects.

OBJECTIVES: The aim of this study was to identify regulatory T cell (Treg) subsets residing in adipose tissue, demonstrate their immunosuppressive functions, and assess the possible role of Sirt1 in their function in overweight subjects. METHODS: Fat samples were obtained by lipoaspiration from healthy overweight (n = 15) and normoweight (n = 11) subjects. We obtained the stromal vascular fraction and then isolated the mononuclear cells by Ficoll-Hypaque sedimentation. The Treg subsets were analyzed by flow cytometry, the expression of Sirt1 and Foxp3 was detected by western blot, and peroxisome proliferator-activated receptor gamma (PPAR-γ) expression was evaluated by qPCR. RESULTS: We detected low numbers of Treg cell subsets displaying the phenotypes CD4+CD25-Foxp3+, CD8+CD25-Foxp3+, and CD4+CD39+Foxp3+ associated with increased body mass index in overweight subjects. We found lower levels of mRNA SIRT1 expression in adipocytes from overweight subjects than in those from normoweight subjects. In contrast, increased amounts of the Sirt1 and Foxp3 proteins in adipose tissue mononuclear cells from overweight subjects were observed. The immunosuppressive function of CD4+CD25+ Treg cells is higher in cells from obese subject than in those from normoweight subject. CONCLUSIONS: Low levels of Treg subsets in overweight subjects with a high percentage of inhibition of proliferation could be related to high levels of the Foxp3 protein. Likewise, the low expression of SIRT1 and PPAR-γ mRNA levels and increased concentration of Sirt1 proteins allows adipose tissue mononuclear cells to respond to stimuli dependent on adenosine receptors and sirtuin pathways.