Carolina Oses

Kupffer cells and hepatocytes maintain liver homeostasis. These cells could be separated based on their size and weight, by centrifugation using a density gradient after a liver perfusion. Here, we describe a methodology to isolate both Kupffer cells and hepatocytes from a single mouse, which provides the unique advantage of studying these two cell types from the same liver.

Obesity and insulin resistance are risk factors for nonalcoholic fatty liver disease (NAFLD), the most common chronic liver disease worldwide. Because no approved medication nor an accurate and noninvasive diagnosis is currently available for NAFLD, there is a clear need to better understand the link between obesity and NAFLD. Lipid accumulation during obesity is known to be associated with oxidative stress and inflammatory activation of liver macrophages (LMs). However, we show that although… Visa mer

Obesity and insulin resistance are risk factors for nonalcoholic fatty liver disease (NAFLD), the most common chronic liver disease worldwide. Because no approved medication nor an accurate and noninvasive diagnosis is currently available for NAFLD, there is a clear need to better understand the link between obesity and NAFLD. Lipid accumulation during obesity is known to be associated with oxidative stress and inflammatory activation of liver macrophages (LMs). However, we show that although LMs do not become proinflammatory during obesity, they display signs of oxidative stress. In livers of both humans and mice, antioxidant nuclear factor erythroid 2–related factor 2 (NRF2) was down-regulated with obesity and insulin resistance, yielding an impaired response to lipid accumulation. At the molecular level, a microRNA-targeting NRF2 protein, miR-144, was elevated in the livers of obese insulin-resistant humans and mice, and specific silencing of miR-144 in murine and human LMs was sufficient to restore NRF2 protein expression and the antioxidant response. These results highlight the pathological role of LMs and their therapeutic potential to restore the impaired endogenous antioxidant response in obesity-associated NAFLD. Visa mindre

Chemopreventive approaches for the treatment of breast cancer have been validated clinically and with in vitro studies. The combined action of tamoxifen/all‐trans retinoic acid was advantageous in MCF‐7 cells, reducing cell proliferation, Bcl‐2, and c‐Myc protein levels and increasing E‐Cadherin protein levels and Gap Junctional Intercellular Communication. We further investigated their combined effect in the presence of bradykinin, a pro‐inflammatory agent, previously reported to contribute to… Visa mer

Chemopreventive approaches for the treatment of breast cancer have been validated clinically and with in vitro studies. The combined action of tamoxifen/all‐trans retinoic acid was advantageous in MCF‐7 cells, reducing cell proliferation, Bcl‐2, and c‐Myc protein levels and increasing E‐Cadherin protein levels and Gap Junctional Intercellular Communication. We further investigated their combined effect in the presence of bradykinin, a pro‐inflammatory agent, previously reported to contribute to the proliferation of breast cancer cells. Bradykinin increased MCF‐7 cell proliferation, c‐Myc levels and ERK1/2 activity. The co‐incubation of bradykinin‐MCF‐7 cells with tamoxifen/all‐trans retinoic acid reduced cell proliferation, ERK1/2 activity, as well as Bcl‐2, c‐Myc, and bradykinin receptor‐2 levels, without altering the enhanced E‐cadherin levels induced by tamoxifen/all‐trans retinoic acid. We showed that the anti‐tumoral effect of tamoxifen/all‐trans retinoic acid is beneficial in MCF‐7 breast cancer cells grown in a bradykinin‐pro‐mitogenic environment, an effect that might be, at least in part, through the MAPK pathway and B2‐bradykinin receptor inhibition. Visa mindre

Diabetic neuropathy (DN) is one of the most frequent and troublesome complications of diabetes mellitus. Evidence from diabetic animal models and diabetic patients suggests that reduced availability of neuroprotective and pro-angiogenic factors in the nerves in combination with a chronic pro-inflammatory microenvironment and high level of oxidative stress, contribute to the pathogenesis of DN. Mesenchymal stem cells (MSCs) are of great interest as therapeutic agents for regenerative purposes… Visa mer

Diabetic neuropathy (DN) is one of the most frequent and troublesome complications of diabetes mellitus. Evidence from diabetic animal models and diabetic patients suggests that reduced availability of neuroprotective and pro-angiogenic factors in the nerves in combination with a chronic pro-inflammatory microenvironment and high level of oxidative stress, contribute to the pathogenesis of DN. Mesenchymal stem cells (MSCs) are of great interest as therapeutic agents for regenerative purposes since they can secrete a broad range of cytoprotective and anti-inflammatory factors. Therefore, the use of the MSC secretome may represent a promising approach for DN treatment. DFX preconditioning of AD-MSCs improves their therapeutic potential and should be considered as a potential strategy for the generation of new alternatives for DN treatment. Visa mindre

The tumor suppressor protein p53 is a transcriptor factor highly mutated in cancer. In the last decades, research has demonstrated that, in addition to its role in the nucleus, p53 has extranuclear functions in the regulation of cellular metabolism, oxidative stress, and drug response. Specifically, p53 has been shown to have a dual role in the regulation of autophagy, a cellular mechanism that allows the turnover of old and damaged proteins and organelles, as well as a key role in cancer… Visa mer

The tumor suppressor protein p53 is a transcriptor factor highly mutated in cancer. In the last decades, research has demonstrated that, in addition to its role in the nucleus, p53 has extranuclear functions in the regulation of cellular metabolism, oxidative stress, and drug response. Specifically, p53 has been shown to have a dual role in the regulation of autophagy, a cellular mechanism that allows the turnover of old and damaged proteins and organelles, as well as a key role in cancer development. Nuclear p53 increases autophagy; however, current research indicates that cytosolic p53, either in wild-type or mutated form, regulate the autophagic pathway independently and in a manner opposite from nuclear p53. In this chapter, we discuss what is known about the nuclear and cytosolic pathways induced by wild-type and p53 mutants in the regulation of autophagy and their impact on tumorigenesis.
Knowledge of the signaling pathways involved in the cytosolic-nuclear interplay will help in the identification of cellular targets that might be used for the development of new cancer therapies. Visa mindre

Understanding how plants sense and respond to changes in nitrogen (N) availability is the first step towards developing strategies for biotechnological applications such as to improve nitrogen-use efficiency. However, components involved in N signaling pathways remain poorly characterized. Calcium is a second messenger in signal transduction pathways in plants and it has been indirectly implicated in nitrate responses. Using aequorin reporter plants we show that nitrate treatments transiently… Visa mer

Understanding how plants sense and respond to changes in nitrogen (N) availability is the first step towards developing strategies for biotechnological applications such as to improve nitrogen-use efficiency. However, components involved in N signaling pathways remain poorly characterized. Calcium is a second messenger in signal transduction pathways in plants and it has been indirectly implicated in nitrate responses. Using aequorin reporter plants we show that nitrate treatments transiently increase cytoplasmic Ca2+ concentration. We found that nitrate also induces cytoplasmic concentration of inositol 1, 4, 5-trisphosphate. Increase in inositol 1, 4, 5-trisphosphate and cytoplasmic Ca2+ levels in response to nitrate treatments was blocked by U732122, a pharmacological inhibitor of phospholipase C, but not by the non-functional phospholipase C inhibitor analog U73343. In addition, increase in cytoplasmic Ca2+ levels in response to nitrate treatments was abolished in mutants of the nitrate transceptor NRT1.1/AtNPF6.3. Gene expression of nitrate-responsive genes was severely affected by pretreatments with Ca2+ channel blockers or phospholipase C inhibitors. These results indicate Ca2+ act as second messenger in the nitrate-signaling pathway of Arabidopsis thaliana. Our results suggest a model where NRT1.1/AtNPF6.3 and a phospholipase C activity mediate the increase of Ca2+ in response to nitrate required for changes in expression of prototypical nitrate-responsive genes. Visa mindre

Prostaglandins (PGs) have been reported to be present in the seminal fluid and cervical mucus, affecting different stages of sperm maturation from spermatogenesis to the acrosome reaction. This study assessed the effects of low physiological PGE2 and PGF2α concentrations on human sperm motility and on the ability of the spermatozoa to bind to the zona pellucida (ZP). Human spermatozoa were isolated from seminal samples with normal concentration and motility parameters and incubated with 1 μM… Visa mer

Prostaglandins (PGs) have been reported to be present in the seminal fluid and cervical mucus, affecting different stages of sperm maturation from spermatogenesis to the acrosome reaction. This study assessed the effects of low physiological PGE2 and PGF2α concentrations on human sperm motility and on the ability of the spermatozoa to bind to the zona pellucida (ZP). Human spermatozoa were isolated from seminal samples with normal concentration and motility parameters and incubated with 1 μM PGE2, 1 μM PGF2α or control solution to determine sperm motility and the ability to bind to human ZP. The effects of both PGs on intracellular calcium levels were determined. Incubation for 2 or 18 h with PGE2 or PGF2α resulted in a significant (P < 0.05) increase in the percentage of spermatozoa with progressive motility. In contrast with PGF2α, PGE2 alone induced an increase in sperm intracellular calcium levels; however, the percentage of sperm bound to the human ZP was doubled for both PGs. These results indicate that incubation of human spermatozoa with low physiological levels of PGE2 or PGF2α increases sperm functions and could improve conditions for assisted reproduction protocols. Visa mindre

Calcium (Ca2+) propagates within tissues serving as an important information carrier. In particular, cilia beat frequency in oviduct cells is partially regulated by Ca2+ changes. Thus, measuring the calcium density and characterizing the traveling wave plays a key role in understanding biological phenomena. However, current methods to measure propagation velocities and other wave characteristics involve several manual or time-consuming procedures. This limits the amount of information that can… Visa mer

Calcium (Ca2+) propagates within tissues serving as an important information carrier. In particular, cilia beat frequency in oviduct cells is partially regulated by Ca2+ changes. Thus, measuring the calcium density and characterizing the traveling wave plays a key role in understanding biological phenomena. However, current methods to measure propagation velocities and other wave characteristics involve several manual or time-consuming procedures. This limits the amount of information that can be extracted, and the statistical quality of the analysis.
Measurements done by different operators showed a high degree of reproducibility. This framework is also extended to a single filter fluorescence experiment, allowing higher sampling rates, and thus an increased accuracy in velocity measurements. Visa mindre