Novel PVA-Hyaluronan-Siloxane Hybrid Nanofiber Mats for Bone Tissue Engineering.

Hyaluronan (HA) is a natural biodegradable biopolymer; its biological functions include cell adhesion, cell proliferation, and differentiation as well as decreasing inflammation, angiogenesis, and regeneration of damaged tissue. This makes it a suitable candidate for fabricating nanomaterials with potential use in tissue engineering. However, HA nanofiber production is restricted due to the high viscosity, low evaporation rate, and high surface tension of HA solutions. Here, hybrids in the form of continuous and randomly aligned polyvinyl alcohol (PVA)-(HA)-siloxane nanofibers were obtained using an electrospinning process. PVA-HA fibers were crosslinked by a 3D siloxane organic-inorganic matrix via sol-gel that restricts natural hydrophilicity and stiffens the structure. The hybrid nanofiber mats were characterized by FT-IR, micro-Raman spectroscopy, SEM, and biological properties. The PVA/HA ratio influenced the morphology of the hybrid nanofibers. Nanofibers with high PVA content (10PVA-8 and 10PVA-10) form mats with few beaded nanofibers, while those with high HA content (5PVA-8 and 5PVA-10) exhibit mats with mound patterns formed by "ribbon-like" nanofibers. The hybrid nanofibers were used as mats to support osteoblast growth, and they showed outstanding biological properties supporting cell adhesion, cell proliferation, and cell differentiation. Importantly, the 5PVA-8 mats show 3D spherical osteoblast morphology; this suggests the formation of tissue growth. These novel HA-based nanomaterials represent a relevant advance in designing nanofibers with unique properties for potential tissue regeneration.

Autophagy as a Target for Non-Immune Intrinsic Functions of Programmed Cell Death-Ligand 1 in Cancer.

Autophagy is a catabolic process that is essential to the maintenance of homeostasis through the cellular recycling of damaged organelles or misfolded proteins, which sustains energy balance. Additionally, autophagy plays a dual role in modulating the development and progression of cancer and inducing a survival strategy in tumoral cells. Programmed cell death-ligand 1 (PD-L1) modulates the immune response and is responsible for maintaining self-tolerance. Because tumor cells exploit the PD-L1-PD-1 interaction to subvert the immune response, immunotherapy has been developed based on the use of PD-L1-blocking antibodies. Recent evidence has suggested a bidirectional regulation between autophagy and PD-L1 molecule expression in tumor cells. Moreover, the research into the intrinsic properties of PD-L1 has highlighted new functions that are advantageous to tumor cells. The relationship between autophagy and PD-L1 is complex and still not fully understood; its effects can be context-dependent and might differ between tumoral cells. This review refines our understanding of the non-immune intrinsic functions of PD-L1 and its potential influence on autophagy, how these could allow the survival of tumor cells, and what this means for the efficacy of anti-PD-L1 therapeutic strategies.

Pfizer-BioNTech vaccine induces the production of cross-reactive antibodies against Trypanosoma cruzi proteins: A preliminary study.

OBJECTIVE: To evaluate the presence of cross-reactivity by anti-severe acute respiratory syndrome coronavirus 2 antibodies induced by the Pfizer-BioNTech vaccine against Trypanosoma cruzi proteins in a screening test. METHODS: Forty-three serum samples were obtained from personnel at the Hospital General Naval de Alta Especialidad in Mexico City who received one or two doses of the vaccine and were tested for T. cruzi infection using four tests: two 'in house' enzyme-linked immunosorbent assays (ELISAs), a commercial ELISA diagnostic kit and an immunoblot test. RESULTS: IgG antibodies against the T. cruzi proteins were present in the serum of unvaccinated subjects and subjects who had received one or two doses of the vaccine. The positivity of the samples against T. cruzi was ruled out by means of a Western Blot assay, where all samples were negative for T. cruzi. CONCLUSION: The data suggest that people convalescing from coronavirus disease 2019 and those who received the Pfizer-BioNTech vaccine exhibit cross-reactive antibodies against T. cruzi antigens in ELISA assays.

Irradiation and parasitism affect the ability of larval hemocytes of Anastrepha obliqua for phagocytosis and the production of reactive oxygen species.

The development of the parasitoid Doryctobracon crawfordi (Viereck) (Hymenoptera: Braconidae) in Anastrepha obliqua (McQuart) (Diptera: Tephritidae) larvae is unviable in nature; however, if the host larva is irradiated at 160 Gy, the parasitoid develops and emerges successfully. This suggests that radiation affects the immune responses of A. obliqua larvae, while the underlying mechanisms remain to be revealed. Using optical and electronic microscopies we determined the number and type of hemocyte populations found inside the A. obliqua larvae, either nonirradiated, irradiated at 160 Gy, parasitized by D. crawfordi, or irradiated and parasitized. Based on flow cytometry, the capacity to produce reactive oxygen species (ROS) was determined by the 123-dihydrorhodamine method in those hemocyte cells. Five cell populations were found in the hemolymph of A. obliqua larvae, two of which (granulocytes and plasmatocytes) can phagocytize and produce ROS. A reduction in the number of cells, mainly of the phagocytic type, was observed, as well as the capacity of these cells to produce ROS, when A. obliqua larvae were irradiated. Both radiation and parasitization decreased the ROS production, and when A. obliqua larvae were irradiated followed by parasitization by D. crawfordi, the reduction of the ROS level was even greater. In contrast, a slight increase in the size of these cells was observed in the hemolymph of the parasitized larvae compared to those in nonparasitized larvae. These results suggest that radiation significantly affects the phagocytic cells of A. obliqua and thus permits the development of the parasitoid D. crawfordi.

Candida glabrata is a successful pathogen: An artist manipulating the immune response.

In recent years, Candida glabrata has emerged as a clinically important pathogen causing invasive infections and death. This review highlights the main virulence factors of this fungus and its strategies to subvert the cell response after invasion. Additionally, we emphasize the intracellular behavior and relationship of C. glabrata with several cells. We contrast these features with some fascinating strategies exhibited by M. tuberculosis to highlight that C. glabrata is a successful pathogen and an artist manipulating the immune response like M. tuberculosis.

CD38 a biomarker and therapeutic target in non-hematopoietic tumors.

The type II transmembrane glycoprotein CD38 has recently been implicated in regulating metabolism and the pathogenesis of multiple conditions, including aging, inflammation and cancer. CD38 is overexpressed in several tumor cells and microenvironment tumoral cells, associated to migration, angiogenesis, cell invasion and progression of the disease. Thus, CD38 has been used as a progression marker for different cancer types as well as in immunotherapy. This review focuses on describing the involvement of CD38 in various non-hematopoietic cancers.

Colostrum IgA1 antibodies recognize antigens from Helicobacter pylori and prevent cytoskeletal changesin human epithelial cells.

Helicobacter pylori is a Gram-negative bacterium found on the luminal surface of the gastric mucosa in at least 50% of the world's human population. The protective effect of breastfeeding against H. pylori infection has been extensively reported; however, the mechanisms behind this protection remain poorly understood. Human IgA from colostrum has reactivity against H. pylori antigens. Despite that IgA1 and IgA2 display structural and functional differences, their reactivity against H. pylori had not been previously determined. We attested titers and reactivity of human colostrum-IgA subclasses by ELISA, immunoblot, and flow cytometry. Colostrum samples from healthy mothers had higher titers of IgA; and IgA1 mostly recognized H. pylori antigens. Moreover, we found a correlation between IgA1 reactivity and their neutralizing effect determined by inhibition of cytoskeletal changes in AGS cells infected with H. pylori. In conclusion, colostrum-IgA reduces H. pylori infection of epithelial gastric cells, suggesting an important role in preventing the bacteria establishment during the first months of life. As a whole, these results suggest that IgA1 from human colostrum provides protection that may help in the development of the mucosal immune system of newborn children.

Ursolic and Oleanolic Acids Induce Mitophagy in A549 Human Lung Cancer Cells.

Ursolic and oleanolic acids are natural isomeric triterpenes known for their anticancer activity. Here, we investigated the effect of triterpenes on the viability of A549 human lung cancer cells and the role of autophagy in their activity. The induction of autophagy, the mitochondrial changes and signaling pathway stimulated by triterpenes were systematically explored by confocal microscopy and western blotting. Ursolic and oleanolic acids induce autophagy in A549 cells. Ursolic acid activates AKT/mTOR pathways and oleanolic acid triggers a pathway independent on AKT. Both acids promote many mitochondrial changes, suggesting that mitochondria are targets of autophagy in a process known as mitophagy. The PINK1/Parkin axis is a pathway usually associated with mitophagy, however, the mitophagy induced by ursolic or oleanolic acid is just dependent on PINK1. Moreover, both acids induce an ROS production. The blockage of autophagy with wortmannin is responsible for a decrease of mitochondrial membrane potential (Δψ) and cell death. The wortmannin treatment causes an over-increase of p62 and Nrf2 proteins promote a detoxifying effect to rescue cells from the death conducted by ROS. In conclusion, the mitophagy and p62 protein play an important function as a survival mechanism in A549 cells and could be target to therapeutic control.

Immune Response to Mucosal Brucella Infection.

Brucellosis is one of the most prevalent bacterial zoonosis of worldwide distribution. The disease is caused by Brucella spp., facultative intracellular pathogens. Brucellosis in animals results in abortion of fetuses, while in humans, it frequently manifests flu-like symptoms and a typical undulant fever, being osteoarthritis a common complication of the chronic infection. The two most common ways to acquire the infection in humans are through the ingestion of contaminated dairy products or by inhalation of contaminated aerosols. Brucella spp. enter the body mainly through the gastrointestinal and respiratory mucosa; however, most studies of immune response to Brucella spp. are performed analyzing models of systemic immunity. It is necessary to better understand the mucosal immune response induced by Brucella infection since this is the main entry site for the bacterium. In this review, some virulence factors and the mechanisms needed for pathogen invasion and persistence are discussed. Furthermore, some aspects of local immune responses induced during Brucella infection will be reviewed. With this knowledge, better vaccines can be designed focused on inducing protective mucosal immune response.

Detection of Brucella abortus by a platform functionalized with protein A and specific antibodies IgG.

Oriented immobilization of antibodies on a sensor surface is critical for enhancing both the antigen-binding capacity and the sensitivity of immunosensors. In this study, we describe a strategy to adsorb immunoglobulin G (IgG) anti-Brucella antibodies onto a silicon surface, oriented by protein A obtained from Staphylococcus aureus (SpA). X-ray photoelectron spectroscopy and atomic force microscopy were used to characterize topographically, morphologically, and chemical changes of the sensor functionalization. The activity of the biosensor was assessed by confocal microscopy, scanning electronic microscopy, and bacteria capture assays (BCA). According to the BCA, the efficiency of Brucella abortus detection with the SpA-IgG anti Brucella biosensor was three-fold higher than that of the random orientated IgG anti Brucella biosensor. The limit of detection was 1 × 106 CFU/ml. These data show that the orientation of antibodies immobilization is crucial to developing immunosensors for bacterial antigen detection as Brucella spp and improve its sensibility level. Functionalization with protein A increases Brucella detection by an antibody-coated surface. Functionalized silicon surface for Brucella detection was characterized by atomic force microscopy, X-ray photoelectron spectroscopy and confocal microscopy.

Candida glabrata survives and replicates in human osteoblasts.

Candida glabrata is an opportunistic pathogen that is considered the second most common cause of candidiasis after Candida albicans Many characteristics of its mechanisms of pathogenicity remain unknown. Recent studies have focused on determining the events that underlie interactions between C. glabrata and immune cells, but the relationship between this yeast and osteoblasts has not been studied in detail. The aim of this study was to determine the mechanisms of interaction between human osteoblasts and C. glabrata, and to identify the roles played by some of the molecules that are produced by these cells in response to infection. We show that C. glabrata adheres to and is internalized by human osteoblasts. Adhesion is independent of opsonization, and internalization depends on the rearrangement of the actin cytoskeleton. We show that C. glabrata survives and replicates in osteoblasts and that this intracellular behavior is related to the level of production of nitric oxide and reactive oxygen species. Opsonized C. glabrata stimulates the production of IL-6, IL-8 and MCP-1 cytokines. Adhesion and internalization of the pathogen and the innate immune response of osteoblasts require viable C. glabrata These results suggest that C. glabrata modulates immunological mechanisms in osteoblasts to survive inside the cell.

Mycobacteria entry and trafficking into endothelial cells.

Endothelial cells are susceptible to infection by mycobacteria, but the endocytic mechanisms that mycobacteria exploit to enter host cells and their mechanisms of intracellular transport are completely unknown. Using pharmacological inhibitors, we determined that the internalization of Mycobacterium tuberculosis (MTB), Mycobacterium smegmatis (MSM), and Mycobacterium abscessus (MAB) is dependent on the cytoskeleton and is differentially inhibited by cytochalasin D, nocodazole, cycloheximide, wortmannin, and amiloride. Using confocal microscopy, we investigated their endosomal trafficking by analyzing Rab5, Rab7, LAMP-1, and cathepsin D. Our results suggest that MSM exploits macropinocytosis to enter endothelial cells and that the vacuoles containing these bacteria fuse with lysosomes. Conversely, the entry of MTB seems to depend on more than one endocytic route, and the observation that only a subset of the intracellular bacilli was associated with phagolysosomes suggests that these bacteria are able to inhibit endosomal maturation to persist intracellularly. The route of entry for MAB depends mainly on microtubules, which suggests that MAB uses a different trafficking pathway. However, MAB is also able to inhibit endosomal maturation and can replicate intracellularly. Together, these findings provide the first evidence that mycobacteria modulate proteins of host endothelial cells to enter and persist within these cells.

Defensin production by human limbo-corneal fibroblasts infected with mycobacteria.

Epithelial cells of the cornea and the conjunctiva constitutively produce antimicrobial peptides; however, the production of defensins by other cell types located around the eye has not been investigated. We analyzed the production of beta-defensins (hBD) and cathelicidin LL-37 during the infection of primary limbo-corneal fibroblasts with M. tuberculosis (MTB), M. abscessus (MAB), and M. smegmatis (MSM). The intracellular survival of each mycobacterium, the production of cytokines and the changes on the distribution of the actin filaments during the infection were also analyzed. Fibroblasts produce basal levels of hBD1 and LL-37 and under PMA stimulation they produce hBD2, hBD3 and overexpress hBD1 and LL-37. MAB induced the highest levels of hBD1 and LL-37 and intermediate levels of IL-6; however, MAB was not eliminated. In addition, MAB induced the greatest change to the distribution of the actin filaments. MTB also produced changes in the structure of the cytoskeleton and induced low levels of hBD1 and IL-6, and intermediate levels of LL-37. The balance of these molecules induced by MTB appeared to contribute to the non-replicative state observed in the limbo-corneal cells. MSM induced the lowest levels of hBD1 and LL-37 but the highest levels of IL-6; MSM was eliminated. The results suggest that mycobacterial infections regulate the production of antimicrobial peptides and cytokines, which in conjunction can contribute to the control of the bacilli.