Reversion by vitamin E treatment of the oxidative damage but not of the advancement in reproductive senescence produced by neonatal hypoxia or hypoxia-ischemia in female rats.

BACKGROUND/AIMS: Few studies address the long-term consequences of perinatal hypoxia (H), a frequent birth complication. Previously we described advanced reproductive senescence (premature loss of regular cyclicity) in female rats subjected to perinatal H or H plus unilateral ischemia (HI) associated with changes in the hypothalamic expression of estrogen and opioid receptors. Our aim is to explore whether hypothalamic inflammation and oxidative damage mediate these reproductive alterations. METHODS: Female rats were subjected on postnatal day (PND) 7 to H (6.5% O2 for 50 min) or HI (H + right carotid artery ligature) and inflammation/oxidative damage markers, such as iNOS, nNOS, insulin-like growth factor (IGF) system expression, glial reaction and macrophage invasion in the medial basal hypothalamus-preoptic area (GFAP Western blot and immunohistochemistry, ED1 immunohistochemistry), were determined. The effect of antioxidant treatment with vitamin E (VE; 1.5 mg/rat on PND 4, 6 and 8) was also explored. RESULTS: No significant cellular inflammatory reactions were observed although GFAP protein was significantly increased at early times after injury. Forty-eight hours after injury iNOS, nNOS and IGF-I mRNA decreased in the HI group, and nNOS in the H group. IGFBP-3 mRNA increased in HI rats at 48 h and 30 days, while it fell at 7 days postinjury in both groups. VE treatment prevented the effects of HI on oxidation/inflammation markers, but did not prevent the premature onset of reproductive senescence or the altered hormone receptors expression. CONCLUSION: These results suggest that the oxidative and inflammatory damage caused by perinatal H or HI may not be responsible for the late-onset reproductive abnormalities.

MSC transplantation: a promising therapeutic strategy to manage the onset and progression of diabetic nephropathy.

Currently, one of the main threats to public health is diabetes mellitus. Its most detrimental complication is diabetic nephropathy (DN), a clinical syndrome associated with kidney damage and an increased risk of cardiovascular disease. Irrespective of the type of diabetes, DN follows a well-known temporal course. The earliest detectable signs are microalbuminuria and histopathological changes including extracellular matrix deposition, glomerular basement membrane thickening, glomerular and mesangial expansion. Later on macroalbuminuria appears, followed by a progressive decline in glomerular filtration rate and the loss of glomerular podocytes, tubulointerstitial fibrosis, glomerulosclerosis and arteriolar hyalinosis. Tight glycemic and hypertension controls remain the key factors for preventing or arresting the progression of DN. Nevertheless, despite considerable educational effort to control the disease, a significant number of patients not only develop DN, but also progress to chronic kidney disease. Therefore, the availability of a strategy aimed to prevent, delay or revert DN would be highly desirable. In this article, we review the pathophysiological features of DN and the therapeutic mechanisms of multipotent mesenchymal stromal cells, also referred to as mesenchymal stem cells (MSCs). The perfect match between them, together with encouraging pre-clinical data available, allow us to support the notion that MSC transplantation is a promising therapeutic strategy to manage DN onset and progression, not only because of the safety of this procedure, but mainly because of the renoprotective potential of MSCs.

Systemic administration of multipotent mesenchymal stromal cells reverts hyperglycemia and prevents nephropathy in type 1 diabetic mice.

Multipotent mesenchymal stromal cells (MSCs), often labeled mesenchymal stem cells, contribute to tissue regeneration in injured bone and cartilage, as well as in the infarcted heart, brain, and kidney. We hypothesize that MSCs might also contribute to pancreas and kidney regeneration in diabetic individuals. Therefore, in streptozotocin (STZ)-induced type 1 diabetes C57BL/6 mice, we tested whether a single intravenous dose of MSCs led to recovery of pancreatic and renal function and structure. When hyperglycemia, glycosuria, massive beta-pancreatic islets destruction, and mild albuminuria were evident (but still without renal histopathologic changes), mice were randomly separated in 2 groups: 1 received 0.5 x 10(6) MSCs that have been ex vivo expanded (and characterized according to their mesenchymal differentiation potential), and the other group received the vehicle. Within a week, only MSC-treated diabetic mice exhibited significant reduction in their blood glucose levels, reaching nearly euglycemic values a month later. Reversion of hyperglycemia and glycosuria remained for 2 months at least. An increase in morphologically normal beta-pancreatic islets was observed only in MSC-treated diabetic mice. Furthermore, in those animals albuminuria was reduced and glomeruli were histologically normal. On the other side, untreated diabetic mice presented glomerular hyalinosis and mesangial expansion. Thus, MSC administration resulted in beta-pancreatic islets regeneration and prevented renal damage in diabetic animals. Our preclinical results suggest bone marrow-derived MSC transplantation as a cell therapy strategy to treat type 1 diabetes and prevent diabetic nephropathy, its main complication.

Advancement of reproductive senescence and changes in the early expression of estrogen, progesterone and micro-opioid receptors induced by neonatal hypoxia in the female rat.

Perinatal hypoxia is a frequent birth complication, and although its early consequences on brain development have been well studied, few studies address any long-term effects. Postnatal insults producing small disturbances in endocrine function can have marked and long-lasting effects. In the present work we studied the effects of two types of perinatal brain injury: global hypoxia (H, 6.5% O2 for 50 min) and hypoxia plus ischemia (HI, ligature of the right carotid artery) on female rat reproductive performance and expression of mediobasal hypothalamus-preoptic area (MBH-PO) estrogen, progesterone and micro-opioid receptors at different times after injury, measuring the mRNA (by semiquantitative RT-PCR) and protein (by Western blot). H or HI advanced approximately 3 months after the appearance of blunted preovulatory LH surges and cyclic irregularities (prolonged estrus) characteristic of the early stages of reproductive senescence. 48 h after H or HI we observed decreases in ERbeta, microOR and PR (only in the H group) mRNAs and in total ER and microOR proteins, followed by increased PR levels (mRNA and protein) 7 days post-injury and by increased microOR protein and ERbeta mRNA in the H group and ERalpha, ERbeta and microOR mRNAs and ER protein in the HI group 30 days post-injury. Thus, an episode of hypoxia suffered during early postnatal life induces premature reproductive senescence on the female rats, accompanied by early changes in some MBH-PO hormone receptors (microOR, ER and PR), whose expression is intimately involved in the regulation of gonadotropin secretion and female sexual cyclicity.

Two complementary strategies to improve cell engraftment in mesenchymal stem cell-based therapy: Increasing transplanted cell resistance and increasing tissue receptivity.

Over the past 2 decades, therapies based on mesenchymal stem cells (MSC) have been tested to treat several types of diseases in clinical studies, due to their potential for tissue repair and regeneration. Currently, MSC-based therapy is considered a biologically safe procedure, with the therapeutic results being very promising. However, the benefits of these therapies are not stable in the long term, and the final outcomes manifest with high inter-patient variability. The major cause of these therapeutic limitations results from the poor engraftment of the transplanted cells. Researchers have developed separate strategies to improve MSC engraftment. One strategy aims at increasing the survival of the transplanted MSCs in the recipient tissue, rendering them more resistant to the hostile microenvironment (cell-preconditioning). Another strategy aims at making the damaged tissue more receptive to the transplanted cells, favoring their interactions (tissue-preconditioning). In this review, we summarize several approaches using these strategies, providing an integral and updated view of the recent developments in MSC-based therapies. In addition, we propose that the combined use of these different conditioning strategies could accelerate the process to translate experimental evidences from pre-clinic studies to the daily clinical practice.

Effects of parity and serum prolactin levels on the incidence and regression of DMBA-induced tumors in OFA hr/hr rats.

Prolactin (PRL) is a key player in the development of mammary cancer. We studied the effects of parity or hyperprolactinemia on mammary carcinogenesis in OFA hr/hr treated with 7,12-dimethylbenzanthracene. They were divided into three groups: nulliparous (Null), primiparous (PL, after pregnancy and lactation), and hyperprolactinemic rats (I, implanted in the arcuate nucleus with 17ß-estradiol). The tumor incidence was similar in the three groups. However, a higher percentage of regressing tumors was evident in the PL group. Serum PRL, mammary development, and mammary ß-casein content were higher in I rats compared to Null. The expression of hormone receptors was similar in the different groups. However, mammary tissue from PL rats bearing tumors had increased expression of PRL and estrogen alpha receptors compared to rats free of tumors. Our results suggest that serum PRL levels do not have relevance on the incidence of tumors, probably because the low levels of PRL in OFA rats are not further decreased by PL like in other strains. However, supraphysiological levels of PRL affect carcinogenesis. PL induces regression of the tumors due to the differentiation produced on the mammary cells. Alterations in the expression of hormonal receptors may be involved in progression and regression of tumors.

Acute sublethal global hypoxia induces transient increase of GAP-43 immunoreactivity in the striatum of neonatal rats.

We assessed immunoreactivity (IR) in the cerebral cortex (CC), hippocampus (Hipp), and striatum (ST) of a growth-associated protein, GAP-43, and of proteins of the synaptic vesicle fusion complex: VAMP-2, Syntaxin-1, and SNAP-25 (SNARE proteins) throughout postnatal development of rats after submitting the animals to acute global postnatal hypoxia (6.5% O(2), 70 min) at postnatal day 4 (PND4). In the CC only the IR of the SNARE protein SNAP-25 increased significantly with age. The hypoxic animals showed the same pattern of IR for SNAP-25, although with lower levels at PND11, and also a significant increase of VAMP-2. SNAP-25 (control): PND11 P < 0.001 vs. PND18, 25, and 40, SNAP-25 (hypoxic): P < 0.001 vs. PND18, 25, and 40; VAMP-2 (hypoxic): P < 0.05 PND11 vs. PND18, and P < 0.01 vs. PND25 and PND40; one-way ANOVA and Bonferroni post-test. In the Hipp, SNAP-25 and syntaxin-1 increased significantly with age, reaching a plateau at PND25 through PND40 in control animals (one-way ANOVA: syntaxin-1: P = 0.043; Bonferroni: NS; SNAP-25: P = 0.013; Bonferroni: P < 0.01 PND11 vs. PND40). Hypoxic rats showed higher levels of significance in the one-way ANOVA than controls (syntaxin-1: P = 0.009; Bonferroni: P < 0.05 PND11 vs. PND25 and P < 0.001 PND11 vs. PND40). In the ST, GAP-43 differed significantly among hypoxic and control animals and the two-way ANOVA revealed significant differences with age (F = 3.23; P = 0.037) and treatment (F = 4.84; P = 0.036). VAMP-2 expression also reached statistical significance when comparing control and treated animals (F = 6.25, P = 0.018) without changes regarding to age. Elevated plus maze test performed at PND40 indicated a lower level of anxiety in the hypoxic animals. At adulthood (12 weeks) learning, memory and locomotor abilities were identical in both groups of animals. With these results, we demonstrate that proteins of the presynaptic structures of the ST are sensitive to acute disruption of homeostatic conditions, such as a temporary decrease of the O(2) concentration. Modifications in the activity of these proteins could contribute to the long term altered responses to stress due to acute hypoxic insult in the neonatal period.

MSC transplantation: a promising therapeutic strategy to manage the onset and progression of diabetic nephropathy

Currently, one of the main threats to public health is diabetes mellitus. Its most detrimental complication is diabetic nephropathy (DN), a clinical syndrome associated with kidney damage and an increased risk of cardiovascular disease. Irrespective of the type of diabetes, DN follows a well-known temporal course. The earliest detectable signs are microalbuminuria and histopathological changes including extracellular matrix deposition, glomerular basement membrane thickening, glomerular and mesangial expansion. Later on macroalbuminuria appears, followed by a progressive decline in glomerular filtration rate and the loss of glomerular podocytes, tubulointerstitial fibrosis, glomerulosclerosis and arteriolar hyalinosis. Tight glycemic and hypertension controls remain the key factors for preventing or arresting the progression of DN. Nevertheless, despite considerable educational effort to control the disease, a significant number of patients not only develop DN, but also progress to chronic kidney disease. Therefore, the availability of a strategy aimed to prevent, delay or revert DN would be highly desirable. In this article, we review the pathophysiological features of DN and the therapeutic mechanisms of multipotent mesenchymal stromal cells, also referred to as mesenchymal stem cells (MSCs). The perfect match between them, together with encouraging pre-clinical data available, allow us to support the notion that MSC transplantation is a promising therapeutic strategy to manage DN onset and progression, not only because of the safety of this procedure, but mainly because of the renoprotective potential of MSCs.

Inflammatory responses of the substantia nigra after acute hypoxia in neonatal rats.

The neocortex and the striatum are the brain regions most known to be particularly vulnerable to acute insults like hypoxia or ischemia. In this work, we assess the possibility of cellular damage to the substantia nigra (SN) after hypoxia-reoxygenation in the new born rat. The aim of the present paper was to evaluate the expression of growth factor IGF-I, and growth factor binding proteins IGFBP-3 and IGFBP-5 genes and induction of NOS family members (nNOS, eNOS and iNOS) and TNF-alpha genes together with glia activation, in the SN at 5 and 48 h after severe hypoxia in the 7 day-old rat, a model for the term human fetus. At early time, while IGFs remain unchanged, we found a transient increase in eNOS and nNOS. Two days after the injury, nNOS expression remained high, iNOS and TNF-alpha increased and also GFAP protein expression was observed together with a profusion of reactive astrocytes distributed throughout the SN. This study on the acute effects of hypoxia on the developing brain provides additional insights into the vulnerability of the SN, a brain region involved in neurodegenerative pathologies.