The effect of progesterone on cardiomyocytes in traumatic brain injury: A randomized study

XV. Steroids

Progesterone: The sexual hormone progesterone is a member of the steroid hormone family, and is the most important representative of the gestagenes sub-group. It plays an elementary role in the female menstruation cycle and is essential for the establishment and the maintenance of a pregnancy, however gestagenes like progesterone are also abundant in males. In 1990, the existence of steroids was described in different cells of the central nervous system (CNS) (Baulieu and Robel, 1990). Up until this point, the effect of sexual hormones on neural cells was rather unknown, other than in the well known regulatory centers of the hypothalamus. Since then the essential enzymes of steroid synthesis, cytochrome P450 side chain cleavage enzyme (P450scc) and 3 β-hydroxysteroid-dehydrogenase (3 β-HSD), have been detected in the central (Mellon et al., 1993) as well as in the peripheral nervous system (Schaeffer et al., 2010). Within the cerebellum Purkinje cells were identified as major sites for neurosteroid formation in the mammalian brain, synthesizing progesterone as well as estradiol (Tsutsui et al., 2011). Traditionally, the effects of progesterone are mediated by genomic mechanisms of classical progesterone receptors which act as transcription factors. Basically, two relevant isoforms, the N-terminal shortened A-form (PR-A, 86 kDa) and the native B-form (PR-B, 110 kDa) are known. Nevertheless, in addition to the genomic signaling pathway, other, non-genomic pathways have been described. The most important member of this non-genomic receptor family seems to be the "progesterone receptor membrane component 1" (PGRMC1). Neural expression of PR-A, PR-B and PGRMC1 could already be proven in different components of the CNS and the peripheral nervous system (PNS) e.g., the hypothalamus, the cerebellum and the dorsal root ganglia (Wessel et al., 2014b). Clinical relevance of progesterone: The effects of neurosteroids like progesterone on neuronal tissue in the CNS and PNS are of enormous therapeutic interest. The clinical relevance of progesterone has already been proven in many studies in different neural glial cells (De Nicola et al., 2013). Indeed, numerous preclinical studies verified the neuroprotective effects of progesterone after cranial traumatic brain and cerebral injuries. Furthermore, experimental data from various animal models emphasize the benefit of progesterone treatment on other neurological disorders like traumatic brain injury, peripheral nerve injury, amyotrophic lateral sclerosis and cerebral ischemia (Wessel et al., 2014b). Progesterone seems to have neuroprotective and anti-inflammatory influences on neuronal cells. For instance, post ischemic treatment with progesterone leads to a reduction of the necrotic area. Based on the versatile application of progesterone, and the outlined positive effects on poor prognosis neurological disorders of the CNS and PNS, the neurosteroids seem to be a very potent group for new therapeutic strategies. But the question is still whether progesterone serves as a universal stimulus for neuronal cells, or if there are therapeutical limitations to a progesterone treatment approach. Is it possible to treat children as well as adults with progesterone after injuries of the CNS or PNS? As there is no answer to this question yet, further basic research is mandatory. Recent studies reviewed the impact of progesterone on neonatal, juvenile and matured cells in the CNS and PNS. In the cerebellum, specifically in rat Purkinje cells, the expression of progesterone with high endogenous concentrations during the neonatal and juvenile periods have been shown (Wessel et al., 2014a). Here, progesterone induces denrito-, spino-, and somatogenesis (Tsutsui et al., 2011; Wessel et al., 2014a). In this study, an age-dependent increase in intracellular progesterone concentrations during the maturation of Purkinje cells and other neurons of the cerebellar cortex, along with an increased receptor expression in juvenile cells suggest that progesterone plays an important role during the physiological development of the cerebellar cortex (Figure 1a). Although Wessel et al. (2014a) demonstrated the expression of the classical progesterone receptors at all developmental stages in rats, the stimulation of matured cells with progesterone had no positive effects concerning neuroplasticity (Figure 1b). Interestingly, at the same time points, the positive impact of progesterone could be verified in the PNS. In primary cultures from chicken dorsal root ganglia (DRG) treated with progesterone, a significant enhancement of neuritic outgrowth was evident (Figure 1a). Blocking of progesterone receptors with mifepristone leads to the extinction of this effect (Olbrich et al., 2013). These results give a strong hint that the use of neurosteroids can be a strategy in pediatric neonatology and traumatology, but at this time point it seems to be limited to juvenile stages and is not applicable in adults. Therefore, we have to investigate and to understand the expression and regulation of the different progesterone receptors in the nervous system, especially in adults. Beside these data progesterone often acts in concert with estrogen. In Purkinje cells, estrogen also promotes dendritic growth, spino- and synaptogenesis during neonatal life (Tsutsui et al., 2011).Figure 1: Impact of progesterone on neuronal cellsa) Progesterone stimulated juvenile neuronal cells in the central nervous system show an increase in dendritogenesis, somatogenesis and spinogenesis. Additionally, cells in the peripheral nervous system show an enlargement in the growth cone after progesterone incubation.b) Adult neurons show no effects after progesterone treatment, neither in the central nervous system nor in the peripheral nervous system. We assume that miRNAs inhibit or degrade the mRNA of progesterone receptors, so that the cell loses its sensitivity to progesterone.c) The regulation of these miRNAs by local or systematic inhibition might increase the number of functional progesterone receptor mRNA molecules. This may result in an increased sensitivity to progesterone in adult stages, possibly leading to effects comparable to those seen in juvenile neuronal cells.MicroRNAs and their relevance in neurology and neurodegenerative diseases: Many studies have been carried out to analyze the function, and subsequently confirm the relevance of microRNAs (miRNA) in the CNS. It has been shown that the biogenesis of miRNAs is crucial for the development and the functionality of neuronal structures. In different independent studies of the cerebral cortex and the cerebellum of mice, it became apparent that inhibition or a complete loss of Dicer leads to different manifestations of neurodegeneration (Hong et al., 2013). MiRNAs are short (21–23 nucleotide, nt), highly conserved, non-coding RNAs. They play a crucial role in posttranscriptional gene regulation e.g., neuroplasticity-related processes (Hommers et al., 2015). The biogenesis of miRNAs, a multistage process, is an important procedure to ensure their functional efficiency. In the first step, the primary transcript of miRNAs (pri-miRNA) is generated in the nucleus. Pri-miRNA has a length of 500–3,000 nucleotides and carries a poly-A-tail at its 3′-end, as well as a 7-methylguanosine cap. Subsequently, the primary transcript is converted into a hairpin structure and cleaved into an approximately 70 nt precursor form (pre-miRNA) by RNase III (Drosha). Pre-miRNA is then transported from the nucleus into the cytoplasm by two proteins, Exportin 5 and Ras-related nuclear protein. In the cytoplasm the RNaseIII, Dicer, and its co-factor Tar RNA-binding protein (TRBP) process the pre-miRNA by cleaving the loop structure and the pre-miRNA into 21–23 nt miRNAs. The single, mature miRNA strands are loaded onto the Argonaute homologue protein (Ago2) in order to form the RNA-induced silencing complex (RISC). In this conformation the miRNA binds to its target mRNA. MiRNAs bind to the 3′ untranslated regions (3′UTR) of their target mRNA and can affect it in two different ways depending on the complementarity to its binding sequence. The transcription can either be inhibited or the mRNA can be degraded completely. A partial complementarity leads to inhibition whereas a perfect base matching causes degradation of the mRNA. Hong et al. (2013) showed that disruption of miRNA biogenesis results in microcephaly in differentiated neurons of the cerebral cortex in Dicer-knockout mice. In comparison to control mice without disruption in pre-miRNA procession, brains of knockout mice were significantly smaller. Total loss of miRNA function leads to a reduced cell soma size of mature neurons and a reduced neurite growth. In a second study it became clear that a knockout of the Dicer enzyme in Purkinje cells is accompanied by dramatic consequences. In contrast to the results of Hong et al., the Dicer-knockout led to cellular death and cerebellar degeneration, and at least induced ataxia (Schaefer et al., 2007). Disruptions due to a knockout of the Dicer enzyme show explicit similarities to different mouse models of neurodegenerative diseases. Apart from the universal step of correct processing of the miRNA, several miRNAs are known to disturb the neuronal development, like the loss of miR-592. Also the involvement of miRNAs in neurodegenerative disease development should not be underestimated. Therefore the emphasis in the investigation of neurodegenerative disorders over the last decade has been concentrated on the involvement of miRNAs. Several miRNAs show a negative effect in the pathogenesis of Parkinson's disease, Alzheimer's disease, Huntington's disease, epilepticus and multiple-system atrophy. For instance in multiple-system atrophy, one single miRNA called miR-202 is the key factor. Effect of miRNAs on progesterone and its receptors: The existence of numerous mRNA targets implicate that miRNAs are capable to regulate thousands of genes. This is why miRNAs are essential in various development stages, tissues and diseases. In terms of progesterone, several studies revealed the effect of miRNAs on progesterone and its receptors. Most of these studies deal with the investigation of miRNAs in breast tumors and their significance in the establishment and maintenance of pregnancy. In both cases the amount of progesterone and its receptors are regulated by different miRNAs. MiR-200a is one key mediator in the decline of progesterone receptor function leading to term and preterm labor (Williams et al., 2012). Progesterone metabolism is up-regulated and the sensitivity of the receptors for progesterone is down-regulated. Apart from these data, progesterone and its receptors could have a strong impact in the nervous system. We know that the classical progesterone receptors are most abundant and sensitive in the early stages of neuronal development. Clinical research in brain injuries, animal models, or even traumata in childhood show promising results when treated with progesterone. The present challenge is to understand the post-transcriptional mechanisms in neuronal cells and to expand the positive effects of progesterone in adulthood. miRNAs, the most important post-transcriptional regulators, are implicated in brain development and in the formation of neurological disorders. Complete comprehension of miRNA function in the neuroscientific field could help to reveal the versatile molecular consequences of miRNA interaction. The understanding of these mechanisms is supposed to be the key to designing new therapeutic tools for the treatment of neuronal damage, in which miRNAs could be used as target molecules for drugs. One promising approach could be the possibility to regulate specific miRNAs by the systematic or local use of miRNA inhibitors, known as antagomirs, and stimulators, known as mimics which are artificial RNA molecules (Figure 1c). An investigation into the gene encoding progesterone resulted in the detection of several binding sites for miRNAs at the 3' UTR which appear to regulate its expression. This opens up new possibilities to interfere with the functionality of the miRNAs that target these sites. One example for a promising miRNA mimic is miR-193b-mimic, a down-regulator of progesterone receptors in a breast cancer cell line (Younger and Corey 2011). This knowledge strongly encourages us to investigate progesterone receptor-regulating miRNAs. Founded on the state of knowledge about the interference of miRNAs in neuronal structures, the main goals are: (1) to reveal regulation mechanisms concerning the classical progesterone receptors, (2) to synthesize mimics and/or antagomirs to replicate the positive impact of progesterone in mature neuronal cells (Figure 1c). We would like to acknowledge D. Terheyden-Keighley for the critical reading of this article. We gratefully thank FoRUM (RUB) for financial support (F812-2014).

Defense and Veterans Brain Injury Center: The First 25 Years

To investigate associations of lifetime history of traumatic brain injury (TBI) with prescription opioid use and misuse among noninstitutionalized adults. Ohio Behavioral Risk Factor Surveillance System (BRFSS) participants in the 2018 cohort who completed the prescription opioid and lifetime history of TBI modules (n = 3448). Secondary analyses of a statewide population-based cross-sectional survey. Self-report of a lifetime history of TBI using an adaptation of the Ohio State University TBI-Identification Method. Self-report of past year: (1) prescription pain medication use (ie, prescription opioid use); and (2) prescription opioid misuse, defined as using opioids more frequently or in higher doses than prescribed and/or using a prescription opioid not prescribed to the respondent. In total, 22.8% of adults in the sample screened positive for a lifetime history of TBI. A quarter (25.5%) reported past year prescription opioid use, and 3.1% met criteria for prescription opioid misuse. A lifetime history of TBI was associated with increased odds of both past year prescription opioid use (adjusted odds ratio [AOR] = 1.52; 95% CI, 1.27-1.83; P < .01) and prescription opioid misuse (AOR = 1.65; 95% CI, 1.08-2.52; P < .05), controlling for sex, age, race/ethnicity, and marital status. Results from this study support the "perfect storm" hypothesis-that persons with a history of TBI are at an increased risk for exposure to prescription opioids and advancing to prescription opioid misuse compared with those without a history of TBI. Routine screening for a lifetime history of TBI may help target efforts to prevent opioid misuse among adults.

Previous studies have demonstrated that progesterone has neuroprotective effects in the central nervous system (CNS) following traumatic brain injury (TBI). Numerous cellular mechanisms have been reported to be important in the neuroprotective effects of progesterone, including the reduction of edema, inflammation and apoptosis, and the inhibition of oxidative stress. However, the effect of progesterone on neuronal protection following TBI remains unclear. The present study aimed to investigate the effects of progesterone on the expression of Nogo-A, an inhibitor of axonal growth, glial fibrillary acidic protein (GFAP), a main component of the glial scar and growth-associated protein-43 (GAP-43), a signaling molecule in neuronal growth in TBI rats. The TBI model was produced by the weight drop method. In total, 75 rats were assigned to three groups: the sham group, TBI group with vehicle treatment and TBI group with progesterone treatment. The protein expression of Nogo-A, GFAP and GAP-43 in the cortex and the hippocampus was examined by immunocytochemistry. TBI rats significantly increased the expression of Nogo-A, GFAP, and GAP-43 at 1, 3, 7 and 14 days post-injury. Progesterone significantly decreased the expression of Nogo-A and GFAP, and upregulated the GAP-43 protein. Our findings suggested that progesterone promotes neuroprotection following TBI by inhibiting the expression of Nogo-A and GFAP, and increasing GAP-43 expression.

The gonadal hormone, progesterone, has been shown to have neuroprotective effects in injured nervous system, including the severity of postinjury cerebral edema. Progesterone's attenuation of edema is accompanied by a sparing of neurons from secondary neuronal death and with improvements in cognitive outcome. In addition, we recently reported that postinjury blood-brain barrier (BBB) leakage, as measured by albumin immunostaining, was significantly lower in progesterone treated than in nontreated rats, supporting a possible protective action of progesterone on the BBB. Because lipid membrane peroxidation is a major contributor to BBB breakdown, we hypothesized that progesterone limits this free radical-induced damage. An antioxidant action, neuroprotective in itself, would also account for progesterone's effects on the BBB, edema, and cell survival after traumatic brain injury. To test progesterone's possible antiperoxidation effect, we compared brain levels of 8-isoprostaglandin F2 alpha (8-isoPGF2 alpha), a marker of lipid peroxidation, 24, 48, and 72 h after cortical contusion in male rats treated with either progesterone or the oil vehicle. The brains of progesterone treated rats contained approximately one-third of the 8-isoPGF2 alpha found in oil-treated rats. These data suggest progesterone has antioxidant effects and support its potential as a treatment for brain injury.

Objective To investigate the effect of progesterone on the expressions of inflammation-related factors of cortical cyclooxygenase-2 ( COX-2 ),prostaglandin E2 ( PGE2 ),inducible nitric oxide synthase (iNOS) and NF-κB in the cortex after traumatic brain injury (TBI) in rats so as to study the possible molecular mechanism of neuroprotective effect of progesterone on TBI.Methods Fortyfive male Spraque-Dawley rats were enrolled in the study and randomly divided into three groups,ie,sham operation group (n =15),TBI group (n =15) and progesterone treatment group (n =15).The rat model of TBI was duplicated with the improved Feeney' s method.The PROG treatment group was given i.p.injections of progesterone ( 16 mg/kg) at 1 and 6 hours after injury.The rats were sacrificed in three groups at 24 hours after injury and the specimens were removed.The changes of the positive cell numbers and protein level of COX-2,PGE2,iNOS and NF-κB in the cortex were examined by immunohistochemistry and Western blot.Results The positive cell numbers and protein levels of COX-2,PGE2,iNOS and NF-κB in the cortex of the TBI group were distinctly higher than those of the sham operation group (P＜O.05).While the positive cell numbers and protein levels of COX-2,PGE2,iNOS and NF-κB in the cortex of the progesterone treatment group were distinctly lower than those of the TBI group ( P ＜O.05).Conclusions Progesterone may exert protective effect on TBI through inhibiting NF-κB activity,blocking the inflammation response course of NF - κB and iNOS and decreasing the expressions of COX-2 and PGE2. Key words: Brain injuries; Progesterone; Inflammation; Nuclear factor-κB

Abstracts from the 3rd Annual Therapeutic Hypothermia and Temperature Management ConferenceMarch 4–5, 2013Miami, FL

Traumatic brain injury (TBI) is a leading cause of death and disability, and the identification of effective, inexpensive and widely practicable treatments for brain injury is of great public health importance worldwide. Progesterone is a naturally produced hormone that has well-defined pharmacokinetics, is widely available, inexpensive, and has steroidal, neuroactive and neurosteroidal actions in the central nervous system. It is, therefore, a potential candidate for treating TBI patients. However, uncertainty exists regarding the efficacy of this treatment. This is an update of our previous review of the same title, published in 2012. To assess the effects of progesterone on neurologic outcome, mortality and disability in patients with acute TBI. To assess the safety of progesterone in patients with acute TBI. We updated our searches of the following databases: the Cochrane Injuries Group's Specialised Register (30 September 2016), the Cochrane Central Register of Controlled Trials (CENTRAL; Issue 9, 2016), MEDLINE (Ovid; 1950 to 30 September 2016), Embase (Ovid; 1980 to 30 September 2016), Web of Science Core Collection: Conference Proceedings Citation Index-Science (CPCI-S; 1990 to 30 September 2016); and trials registries: Clinicaltrials.gov (30 September 2016) and the World Health Organization (WHO) International Clinical Trials Registry Platform (30 September 2016). We included randomised controlled trials (RCTs) of progesterone versus no progesterone (or placebo) for the treatment of people with acute TBI. Two review authors screened search results independently to identify potentially relevant studies for inclusion. Independently, two review authors selected trials that met the inclusion criteria from the results of the screened searches, with no disagreement. We included five RCTs in the review, with a total of 2392 participants. We assessed one trial to be at low risk of bias; two at unclear risk of bias (in one multicentred trial the possibility of centre effects was unclear, whilst the other trial was stopped early), and two at high risk of bias, due to issues with blinding and selective reporting of outcome data.All included studies reported the effects of progesterone on mortality and disability. Low quality evidence revealed no evidence of a difference in overall mortality between the progesterone group and placebo group (RR 0.91, 95% CI 0.65 to 1.28, I² = 62%; 5 studies, 2392 participants, 2376 pooled for analysis). Using the GRADE criteria, we assessed the quality of the evidence as low, due to the substantial inconsistency across studies.There was also no evidence of a difference in disability (unfavourable outcomes as assessed by the Glasgow Outcome Score) between the progesterone group and placebo group (RR 0.98, 95% CI 0.89 to 1.06, I² = 37%; 4 studies; 2336 participants, 2260 pooled for analysis). We assessed the quality of this evidence to be moderate, due to inconsistency across studies.Data were not available for meta-analysis for the outcomes of mean intracranial pressure, blood pressure, body temperature or adverse events. However, data from three studies showed no difference in mean intracranial pressure between the groups. Data from another study showed no evidence of a difference in blood pressure or body temperature between the progesterone and placebo groups, although there was evidence that intravenous progesterone infusion increased the frequency of phlebitis (882 participants). There was no evidence of a difference in the rate of other adverse events between progesterone treatment and placebo in the other three studies that reported on adverse events. This updated review did not find evidence that progesterone could reduce mortality or disability in patients with TBI. However, concerns regarding inconsistency (heterogeneity among participants and the intervention used) across included studies reduce our confidence in these results.There is no evidence from the available data that progesterone therapy results in more adverse events than placebo, aside from evidence from a single study of an increase in phlebitis (in the case of intravascular progesterone).There were not enough data on the effects of progesterone therapy for our other outcomes of interest (intracranial pressure, blood pressure, body temperature) for us to be able to draw firm conclusions.Future trials would benefit from a more precise classification of TBI and attempts to optimise progesterone dosage and scheduling.

Using the NGF/IL-6 ratio as a reliable criterion to show the beneficial effects of progesterone after experimental diffuse brain injury

Neuropsychiatric Complications of Traumatic Brain Injury: A Critical Review of the Literature (A Report by the ANPA Committee on Research)

The Neuroendocrine Effects of Traumatic Brain Injury

Neuroendocrine dysfunction after traumatic brain injury (TBI) is under-diagnosed, under-treated, and may adversely affect the rate of recovery. Single or multiple pituitary-target hormone disruption occurs in up to two-thirds of persons with TBI, most commonly affecting the gonadal and growth hormone axes. The time course of decline in and recovery of pituitary function in relation to cognitive dysfunction and rehabilitation progress are not well described. This article reviews the clinical spectrum of neuroendocrine deficits after TBI and their underlying mechanisms. Future studies of the effects of hormonal replacement on recovery are recommended.

To investigate the impact of traumatic injury on the developing prefrontal-temporal adolescent cortex, and correlated brain structural measures with neurocognitive functioning. Nineteen adolescents (12 males, 7 females, age range: 11-17y, mean 15y 8mo, standard deviation 1y 7mo, median 15y 11mo) with traumatic brain injury (TBI) were included. Cortical thickness of frontal and temporal lobes was assessed using magnetic resonance imaging. We correlated cortical thickness of prefrontal-temporal regions with age, time since injury, and neurocognitive functioning, and compared these results with a matched control cohort without TBI. We found thinner prefrontal (p=0.039) and temporal cortices (p=0.002) in adolescents with TBI compared to typically developing children. Furthermore, significant age effect was observed on the prefrontal (r=-0.75, p=0.003) and temporal (r=-0.66, p=0.013) cortical thickness in typically developing adolescents, but not in adolescents with TBI. Executive function (measured using the Behaviour Rating Inventory of Executive Function questionnaire, with lower scores meaning higher functioning) was correlated with prefrontal cortical thickness in typically developing adolescents (r=0.72, p=0.009). Opposite trends were found for correlations between cortical thickness and executive function in the TBI and control cohort. Structural maturation in typically developing adolescents correlates with functional development: the older the adolescent, the thinner the prefrontal cortex, the better executive function. In adolescents with TBI we observed an opposite trend, that appeared significantly different from the control group: the thinner the prefrontal and temporal cortex, the worse executive functioning. Cortical thickness is negatively correlated with age in typically developing adolescents. Prefrontal cortex thickness correlates negatively with executive function in typically developing adolescents. Correlations between cortical thickness and executive functioning rise for adolescents without traumatic brain injury (TBI). Correlations between cortical thickness and executive functioning fall for adolescents with TBI. Adolescents with TBI have a long-term impairment of adaptive functioning in daily living.

Personal Bankruptcy After Traumatic Brain or Spinal Cord Injury: The Role of Medical Debt