Descriptions of Childhood Trauma, Effects of the Trauma, and How Adults Moved Through the Trauma to Normalized Behavior

Problem. People who are not able to overcome the effects of childhood trauma often waste their human potential on substance abuse and illegal and illicit lifestyles; they end up in prison, uneducated, or develop self-destructive behavior. They often struggle with poor learning and social skills and are not able to be successful in life. However, some individuals do succeed. This study interviewed eight individuals who suffered childhood trauma and developed ways of overcoming that trauma to live prosperous, productive, and, in many regards, full lives. Method. This qualitative study used interviews to explore how adults traumatized as children described childhood trauma and its effect, and their ability to move through the trauma and live normalized adult lives. The eight individuals were located by personal contacts and snowball referrals. Face-to-face interviews took place in public and private locations chosen by individuals. The eight participants consisted of four individuals and two couples. The open-ended questions during the interviews allowed each participant to comfortably disclose their sensitive stories of their past traumatic experiences and how they eventually were supported, encouraged, and nurtured to normalized adult behavior. The writing process began with transcription of recorded interviews. The first step of the data analysis process was to organize the data into details and then look at the individual pieces of information as a whole. A precoding process was used to identify similarities and differences in interviews. The following code systems were applied in the first cycle of coding: (a) in vivo codes: taken directly from what participants said, (b) descriptive codes: summarize the primary topic, usually a noun, (c) process codes: words or phrases that capture action, (d) values codes: assess participants’ integrated value, attitude, and belief systems at work, and (e) emotion codes: describe a participant’s emotional experience, primary emotions, occurring with specific experience or period of time. The idea was to look for coding patterns. Pseudonyms were used to maintain anonymity and confidentiality. This study also used Dr. Lenore Terr’s theory of childhood trauma that concludes that childhood trauma has four lasting characteristics: visualized memories, repetitive behaviors, trauma-specific fears, and changed attitudes about people, life, and the future. The stories in this study were written to reveal and celebrate each individual’s success as they moved through childhood trauma to normalized behavior. The interviewees of four individuals and two sets of couples were of various ages ranging from their early 20s to a gentleman in his mid-60s. Participants varied in race and socioeconomic status. It was important that the couples’ stories be meshed and joined together, as their lives and journeys are incomplete without each other’s interpretation of their individual and coupled transitions through childhood trauma to normalized adult behavior. Some shared details of their trauma. Others gave brief overviews of their abuse but detailed their tumultuous and eventual resilient journeys into adulthood. Chapter 4 is divided up into six sections of the four individuals and two couples who were interviewed. Each section contains six subsections. The first subsection is a brief introduction with participants’ background information. The significant themes include: (a) family systems and childhood trauma, (b) feelings about the trauma, (c) results of abuse and trauma, (d) coping with support, and (e) resiliency within the process of being rescued. Results. Cross-case analysis revealed repetitive patterns and themes, which corresponded to the research question: How do adults traumatized as children describe childhood trauma, effects of the trauma, and their ability to move through the trauma to normalized behavior? Themes that emerged from cross-case analyses and a developmental timeline were: (a) reported childhood trauma, (b) effects of trauma, (c) main childhood caregiver, (d) support systems reported, and (e) evidence of resiliency. The reason for choosing these themes was that each interview revealed similarities. Each individual reported various traumas that affected their behavior and emotions. Each individual had a main childhood caregiver and support system, though the support system was not necessarily the main caregiver. All gave evidence of resiliency. The effects described by the eight participants coincide with Lenore Terr’s theory of childhood trauma mentioned above. Each story and trauma is also different. Neglect, abandonment, poverty, and substance and physical abuse are observable offenders. But when a child does not know how to speak up and report secret sexual assault or emotional abuse, the trauma becomes less obvious. All of the stories must be told and voices heard. Nevertheless, children of trauma can heal. Participants in this study stated they were on the journey towards recovery from childhood trauma to normalized adult behavior. Conclusion. The themes addressed in this study can be interconnected and related to the research question: How do adults traumatized as children describe childhood trauma, effects of the trauma, and their ability to move through the trauma to normalized behavior? The themes that emerged from the stories were: (a) reported childhood trauma, (b) effects of childhood trauma, (c) main childhood caregiver, (d) support systems reported, and (e) evidence of resiliency. Each adult interviewed volunteered and was willing to describe his or her childhood trauma. For example, neglect, which included divorce, poverty, substance abuse, and violence, was the most prevalent of traumas. Subsequent was behavioral and emotional trauma. All participants told of childhood trauma that affected cognitive and social development. The majority of the trauma was caused and inflicted by caregivers due to neglect. It is unknown if there was intergenerational trauma or if caregiver stress was the initiator of the childhood trauma. Substance abuse of caregivers was also described as a baseline for familial stress. Most support systems were not available to participants as children. These systems include: (a) psychological and psychiatric counseling, (b) education and social, (c) familial and community, (d) and spiritual. Internal and self-directed support and motivation were inferred by all participants, which were fueled by love, faith, and encouragement from external support to motivate participants to normalized adult behavior. Evidence of resiliency was told by each individual. Compassionate giving back to communities was major evidence of healing as well as reported self-respect, hard working in family and community responsibilities, maturity, forgiveness, and security

High incidence of Noonan syndrome features including short stature and pulmonic stenosis in patients carrying NF1 missense mutations affecting p.Arg1809: genotype-phenotype correlation

Neurofibromatosis type 1 (NF1) is one of the most frequent genetic disorders, affecting 1:3,000 worldwide. Identification of genotype-phenotype correlations is challenging because of the wide range clinical variability, the progressive nature of the disorder, and extreme diversity of the mutational spectrum. We report 136 individuals with a distinct phenotype carrying one of five different NF1 missense mutations affecting p.Arg1809. Patients presented with multiple cafe-au-lait macules (CALM) with or without freckling and Lisch nodules, but no externally visible plexiform neurofibromas or clear cutaneous neurofibromas were found. About 25% of the individuals had Noonan-like features. Pulmonic stenosis and short stature were significantly more prevalent compared with classic cohorts (P<0.0001). Developmental delays and/or learning disabilities were reported in over 50% of patients. Melanocytes cultured from a CALM in a segmental NF1-patient showed two different somatic NF1 mutations, p.Arg1809Cys and a multi-exon deletion, providing genetic evidence that p.Arg1809Cys is a loss-of-function mutation in the melanocytes and causes a pigmentary phenotype. Constitutional missense mutations at p.Arg1809 affect 1.23% of unrelated NF1 probands in the UAB cohort, therefore this specific NF1 genotype-phenotype correlation will affect counseling and management of a significant number of patients

Impact of glucocorticoid receptor density on ligand-independent dimerization, cooperative ligand-binding and basal priming of transactivation: a cell culture model

Glucocorticoid receptor (GR) levels vary between tissues and individuals and are altered by physiological and pharmacological effectors. However, the effects and implications of differences in GR concentration have not been fully elucidated. Using three statistically different GR concentrations in transiently transfected COS-1 cells, we demonstrate, using co-immunoprecipitation (CoIP) and fluorescent resonance energy transfer (FRET), that high levels of wild type GR (wtGR), but not of dimerization deficient GR (GRdim), display ligand-independent dimerization. Whole-cell saturation ligand-binding experiments furthermore establish that positive cooperative ligand-binding, with a concomitant increased ligand-binding affinity, is facilitated by ligand-independent dimerization at high concentrations of wtGR, but not GRdim. The down-stream consequences of ligand-independent dimerization at high concentrations of wtGR, but not GRdim, are shown to include basal priming of the system as witnessed by ligand-independent transactivation of both a GRE-containing promoter-reporter and the endogenous glucocorticoid (GC)-responsive gene, GILZ, as well as ligand-independent loading of GR onto the GILZ promoter. Pursuant to the basal priming of the system, addition of ligand results in a significantly greater modulation of transactivation potency than would be expected solely from the increase in ligand-binding affinity. Thus ligand-independent dimerization of the GR at high concentrations primes the system, through ligand-independent DNA loading and transactivation, which together with positive cooperative ligand-binding increases the potency of GR agonists and shifts the bio-character of partial GR agonists. Clearly GR-levels are a major factor in determining the sensitivity to GCs and a critical factor regulating transcriptional programs

Robust estimation of bacterial cell count from optical density

Optical density (OD) is widely used to estimate the density of cells in liquid culture, but cannot be compared between instruments without a standardized calibration protocol and is challenging to relate to actual cell count. We address this with an interlaboratory study comparing three simple, low-cost, and highly accessible OD calibration protocols across 244 laboratories, applied to eight strains of constitutive GFP-expressing E. coli. Based on our results, we recommend calibrating OD to estimated cell count using serial dilution of silica microspheres, which produces highly precise calibration (95.5% of residuals &lt;1.2-fold), is easily assessed for quality control, also assesses instrument effective linear range, and can be combined with fluorescence calibration to obtain units of Molecules of Equivalent Fluorescein (MEFL) per cell, allowing direct comparison and data fusion with flow cytometry measurements: in our study, fluorescence per cell measurements showed only a 1.07-fold mean difference between plate reader and flow cytometry data

Effect of angiotensin-converting enzyme inhibitor and angiotensin receptor blocker initiation on organ support-free days in patients hospitalized with COVID-19

IMPORTANCE Overactivation of the renin-angiotensin system (RAS) may contribute to poor clinical outcomes in patients with COVID-19. Objective To determine whether angiotensin-converting enzyme (ACE) inhibitor or angiotensin receptor blocker (ARB) initiation improves outcomes in patients hospitalized for COVID-19. DESIGN, SETTING, AND PARTICIPANTS In an ongoing, adaptive platform randomized clinical trial, 721 critically ill and 58 non–critically ill hospitalized adults were randomized to receive an RAS inhibitor or control between March 16, 2021, and February 25, 2022, at 69 sites in 7 countries (final follow-up on June 1, 2022). INTERVENTIONS Patients were randomized to receive open-label initiation of an ACE inhibitor (n = 257), ARB (n = 248), ARB in combination with DMX-200 (a chemokine receptor-2 inhibitor; n = 10), or no RAS inhibitor (control; n = 264) for up to 10 days. MAIN OUTCOMES AND MEASURES The primary outcome was organ support–free days, a composite of hospital survival and days alive without cardiovascular or respiratory organ support through 21 days. The primary analysis was a bayesian cumulative logistic model. Odds ratios (ORs) greater than 1 represent improved outcomes. RESULTS On February 25, 2022, enrollment was discontinued due to safety concerns. Among 679 critically ill patients with available primary outcome data, the median age was 56 years and 239 participants (35.2%) were women. Median (IQR) organ support–free days among critically ill patients was 10 (–1 to 16) in the ACE inhibitor group (n = 231), 8 (–1 to 17) in the ARB group (n = 217), and 12 (0 to 17) in the control group (n = 231) (median adjusted odds ratios of 0.77 [95% bayesian credible interval, 0.58-1.06] for improvement for ACE inhibitor and 0.76 [95% credible interval, 0.56-1.05] for ARB compared with control). The posterior probabilities that ACE inhibitors and ARBs worsened organ support–free days compared with control were 94.9% and 95.4%, respectively. Hospital survival occurred in 166 of 231 critically ill participants (71.9%) in the ACE inhibitor group, 152 of 217 (70.0%) in the ARB group, and 182 of 231 (78.8%) in the control group (posterior probabilities that ACE inhibitor and ARB worsened hospital survival compared with control were 95.3% and 98.1%, respectively). CONCLUSIONS AND RELEVANCE In this trial, among critically ill adults with COVID-19, initiation of an ACE inhibitor or ARB did not improve, and likely worsened, clinical outcomes. TRIAL REGISTRATION ClinicalTrials.gov Identifier: NCT0273570

The β2_{2}-Subunit of Voltage-Gated Calcium Channels Regulates Cardiomyocyte Hypertrophy

L-type voltage-gated calcium channels (LTCCs) regulate crucial physiological processes in the heart. They are composed of the Cavα1 pore-forming subunit and the accessory subunits Cavβ, Cavα2δ, and Cavγ. Cavβ is a cytosolic protein that regulates channel trafficking and activity, but it also exerts other LTCC-independent functions. Cardiac hypertrophy, a relevant risk factor for the development of congestive heart failure, depends on the activation of calcium-dependent pro-hypertrophic signaling cascades. Here, by using shRNA-mediated Cavβ silencing, we demonstrate that Cavβ2 downregulation enhances α1-adrenergic receptor agonist-induced cardiomyocyte hypertrophy. We report that a pool of Cavβ2 is targeted to the nucleus in cardiomyocytes and that the expression of this nuclear fraction decreases during in vitro and in vivo induction of cardiac hypertrophy. Moreover, the overexpression of nucleus-targeted Cavβ2 in cardiomyocytes inhibits in vitro-induced hypertrophy. Quantitative proteomic analyses showed that Cavβ2 knockdown leads to changes in the expression of diverse myocyte proteins, including reduction of calpastatin, an endogenous inhibitor of the calcium-dependent protease calpain. Accordingly, Cavβ2-downregulated cardiomyocytes had a 2-fold increase in calpain activity as compared to control cells. Furthermore, inhibition of calpain activity in Cavβ2-downregulated cells abolished the enhanced α1-adrenergic receptor agonist-induced hypertrophy observed in these cells. Our findings indicate that in cardiomyocytes, a nuclear pool of Cavβ2 participates in cellular functions that are independent of LTCC activity. They also indicate that a downregulation of nuclear Cavβ2 during cardiomyocyte hypertrophy promotes the activation of calpain-dependent hypertrophic pathways

TRPC4α and TRPC4β Similarly Affect Neonatal Cardiomyocyte Survival during Chronic GPCR Stimulation

<div><p>The Transient Receptor Potential Channel Subunit 4 (TRPC4) has been considered as a crucial Ca²⁺ component in cardiomyocytes promoting structural and functional remodeling in the course of pathological cardiac hypertrophy. TRPC4 assembles as homo or hetero-tetramer in the plasma membrane, allowing a non-selective Na⁺ and Ca²⁺ influx. Gαq protein-coupled receptor (GPCR) stimulation is known to increase TRPC4 channel activity and a TRPC4-mediated Ca²⁺ influx which has been regarded as ideal Ca²⁺ source for calcineurin and subsequent nuclear factor of activated T-cells (NFAT) activation. Functional properties of TRPC4 are also based on the expression of the TRPC4 splice variants TRPC4α and TRPC4β. Aim of the present study was to analyze cytosolic Ca²⁺ signals, signaling, hypertrophy and vitality of cardiomyocytes in dependence on the expression level of either TRPC4α or TRPC4β. The analysis of Ca²⁺ transients in neonatal rat cardiomyocytes (NRCs) showed that TRPC4α and TRPC4β affected Ca²⁺ cycling in beating cardiomyocytes with both splice variants inducing an elevation of the Ca²⁺ transient amplitude at baseline and TRPC4β increasing the Ca²⁺ peak during angiotensin II (Ang II) stimulation. NRCs infected with TRPC4β (Ad-C4β) also responded with a sustained Ca²⁺ influx when treated with Ang II under non-pacing conditions. Consistent with the Ca²⁺ data, NRCs infected with TRPC4α (Ad-C4α) showed an elevated calcineurin/NFAT activity and a baseline hypertrophic phenotype but did not further develop hypertrophy during chronic Ang II/phenylephrine stimulation. Down-regulation of endogenous TRPC4α reversed these effects, resulting in less hypertrophy of NRCs at baseline but a markedly increased hypertrophic enlargement after chronic agonist stimulation. Ad-C4β NRCs did not exhibit baseline calcineurin/NFAT activity or hypertrophy but responded with an increased calcineurin/NFAT activity after GPCR stimulation. However, this effect was not translated into an increased propensity towards hypertrophy but rather less hypertrophy during GPCR stimulation. Further analyses revealed that, although hypertrophy was preserved in Ad-C4α NRCs and even attenuated in Ad-C4β NRCs, cardiomyocytes had an increased apoptosis rate and thus were less viable after chronic GPCR stimulation. These findings suggest that TRPC4α and TRPC4β differentially affect Ca²⁺ signals, calcineurin/NFAT signaling and hypertrophy but similarly impair cardiomyocyte viability during GPCR stimulation.</p></div

TRPC4α and TRPC4β lower cardiomyocyte viability after chronic agonist stimulation.

<p>The vitality of cardiomyocytes expressing βgal (Ad- βgal), TRPC4α (Ad-C4α) or TRPC4β (Ad-C4β) was determined after angiotensin II/phenylephrine (Ang II/PE) or vehicle (CON) treatment for 24 h by applying the trypan blue exclusion assay. The assay was performed in duplicate and average values were quantified relative to Ad-βgal CON; n = 3 experiments. *<i>P</i><0.05 vs Ad-C4α and Ad-C4β Ang II/PE. B. ^#<i>P</i><0.05 vs Ad-C4α and Ad-C4β CON.</p

Increased apoptotic rate in cardiomyocytes overexpressing TRPC4α or TRPC4β.

<p>A, Cleaved caspase-3 expression levels were detected in neonatal rat cardiomyocytes (NRCs) after Ang II/PE or vehicle (CON) treatment. GAPDH: loading control. Myc: detection of TRPC4α and TRPC4β. Shown are representative immunoblots from three independent experiments. B, Average quantified values of cleaved caspase-3 expression levels relative to Ad-βgal CON for n = 3 experiments. C, Capase-3 activity was compared between Ad-βgal, Ad-TRPC4α and Ad-TRPC4β in NRCs. Caspase-3 assays were performed in duplicate and average values were quantified relative to Ad-βgal CON; n = 3 experiments.</p

NFATc1 nuclear translocation is promoted by TRPC4α at baseline and by TRPC4β upon agonist stimulation.

<p>A, Neonatal rat cardiomyocytes (NRCs) were co-infected with Ad-βgal, Ad-TRPC4α or Ad-TRPC4β/ NFATc1-GFP. Nuclear localization of NFATc1-GFP was used as an indicator for its activation. Green: NFATc1-GFP. Scale bar: 20 μm. B, Quantification of NFATc1 localized in the nucleus of NRCs that were stimulated with angiotensin II/phenylephrine (Ang II/PE; 1 μM/50 μM) or vehicle (CON). *<i>P</i><0.05 vs unstimulated; ^#<i>P</i><0.05 vs Ad-TRPC4α or Ad-TRPC4β CON, respectively; ^$<i>P</i><0.05 vs Ad-βgal and Ad-TRPC4α Ang II/PE. Cells from at least three different experiments were analyzed. N-number of cells is indicated in the bars.</p