Racial and ethnic differences in diagnosis age and blood biomarkers in a pediatric inflammatory bowel disease cohort.

OBJECTIVE: Prompt diagnosis of pediatric-onset inflammatory bowel disease (IBD) is crucial for preventing a complicated disease course; however, it is not well understood how social determinants of health might affect pediatric IBD diagnosis. This study examined differences in diagnosis age, biomarkers of disease severity, and anthropometrics with sociodemographic factors in a pediatric IBD cohort. METHODS: Pediatric IBD patients (n = 114) and their parents/caregivers were enrolled from the Children's of Alabama Pediatric IBD Clinic in Birmingham, Alabama. Primary analyses examined associations of child race and ethnicity, parental income, parental education, single-parent household status, insurance type, and distance to a tertiary pediatric gastroenterology referral center with diagnosis age. Secondary analyses examined differences in biomarker levels, height, and body mass index at the time of diagnosis. RESULTS: Racial and ethnic minority children were diagnosed at an older age compared to Non-Hispanic White children (14.4 ± 0.40 vs. 11.7 ± 0.38 years; p < 0.001), and this trend was robust to adjustment with other sociodemographic variables. Parental attainment of a college education attenuated the link between minority race and ethnicity and the likelihood of older age at diagnosis, while other sociodemographic variables had no moderating effect. Racial and ethnic minority children were 5.7 times more likely to have clinically elevated erythrocyte sedimentation rate at diagnosis compared to Non-Hispanic White children (p = .024). CONCLUSIONS: These results suggest that child race and ethnicity may exert a primary effect on the age at diagnosis with pediatric-onset IBD. This study highlights the need for further research on racial and ethnic disparities to promote health equity in pediatric-onset IBD.

Early Life Stress in Mice Leads to Impaired Colonic Corticosterone Production and Prolonged Inflammation Following Induction of Colitis.

BACKGROUND: Early life stress (ELS) is an environmental trigger believed to promote increased risk of IBD. Our goal was to identify mechanisms whereby ELS in mice affects susceptibility to and/or severity of gut inflammation. METHODS: We utilized 2 published animal models of ELS. In the first model, newborn mice were separated from the dam daily for 4 to 8 hours starting on postnatal day 2 and then weaned early on postnatal day 17. Control mice were left undisturbed with the dams until weaning on postnatal day 21. In the second model, dams were fed dexamethasone or vehicle ad libitum in drinking water on postpartum days 1 to 14. Plasma and colonic corticosterone were measured in juvenile and adult mice. Colitis was induced in 4-week-old mice via intraperitoneal injection of interleukin (IL)-10 receptor blocking antibody every 5 days for 15 days. Five or 15 days later, colitis scores and transcripts for Tnf, glucocorticoid receptors, and steroidogenic enzymes were measured. RESULTS: Mice exposed to ELS displayed reduced plasma and colonic corticosterone. Control animals showed improvements in indices of inflammation following cessation of interleukin-10 receptor blockade, whereas ELS-exposed animals maintained high levels of Tnf and histological signs of colitis. In colitic animals, prior exposure to ELS was associated with significantly lower expression of genes associated with corticosterone synthesis and responsiveness. Finally, TNF stimulation of colonic crypt cells from ELS mice led to increased inhibition of corticosterone synthesis. CONCLUSIONS: Our study identifies impaired local glucocorticoid production and responsiveness as a potential mechanism whereby ELS predisposes to chronic colitis in susceptible hosts.

Using 2 distinct animal models, this study shows that in mice, early life stress leads to reduced colonic corticosterone and that induction of colitis after stress removal results in reduced transcription of glucocorticoid synthesis genes, increased Tnf, and enhanced chronicity of intestinal inflammation.

A continuing clinical education course to maintain clinical competencies and foster new clinical knowledge during the graduate school years of MD-PhD training.

Introduction: Most students in MD-PhD programs take a leave of absence from medical school to complete PhD training, which promotes a natural loss of clinical skills and knowledge and could negatively impact a student's long-term clinical knowledge. To address this concern, clinical refresher courses in the final year of PhD training have traditionally been used; however, effectiveness of such courses versus a longitudinal clinical course spanning all PhD training years is unclear. Methods: The University of Alabama at Birmingham MD-PhD Program implemented a comprehensive continuing clinical education (CCE) course spanning PhD training years that features three course components: (1) clinical skills; (2) clinical knowledge; and (3) specialty exposure activities. To evaluate course effectiveness, data from an anonymous student survey completed at the end of each semester were analyzed. Results: Five hundred and ninety-seven surveys were completed by MD-PhD students from fall 2014 to 2022. Survey responses indicated that the majority of students found the course helpful to: maintain clinical skills and knowledge (544/597, 91% and 559/597, 94%; respectively), gain exposure to clinical specialties (568/597, 95%), and prepare them for responsibilities during clinical clerkships. During semesters following lockdowns from the COVID-19 pandemic, there were significant drops in students' perceived preparedness. Conclusions: Positive student survey feedback and improved preparedness to return to clinic after development of the course suggests the CCE course is a useful approach to maintain clinical knowledge during research training.

FVB/N mouse strain regulatory T cells differ in phenotype and function from the C57BL/6 and BALB/C strains.

Regulatory T cells (Treg) are vital to the maintenance of immune homeostasis. The genetic background of an inbred mouse strain can have a profound effect on the immune response in the animal, including Treg responses. Most Treg studies focus on animals created on the C57BL/6 or BALB/c background. Recent studies have demonstrated a difference in the phenotype and behavior of C57BL/6 and BALB/c Tregs. In this study, we have investigated the function of FVB/N Tregs compared to C57BL/6 and BALB/c. We observed that while FVB/N Tregs appear to suppress normally in a cell contact-dependent system, FVB/N Tregs are less capable of suppressing when regulation depends on the secretion of a soluble factor. FVB/N Tregs produce IL-10; however, TGF-ß was not detected in any culture from C57BL/6 or FVB/N. C57BL/6 Foxp3+ Tregs expressed more of the TGF-ß-related proteins glycoprotein-A repetitions predominant (GARP) and latency-associated peptide (LAP) on the cell surface than both FVB/N and BALB/c, but C57BL/6 Tregs expressed significantly less Ctse (Cathepsin E) mRNA. Each strain displayed different abilities of thymic Tregs (tTreg) to maintain Foxp3 expression and had a varying generation of induced Tregs (iTregs). In vitro generated FVB/N iTregs expressed significantly less GARP and LAP. These results suggest Tregs of different strains have varying phenotypes and dominant mechanisms of action for the suppression of an immune response. This information should be taken into consideration when Tregs are examined in future studies, particularly for therapeutic purposes in a genetically diverse population.

A dimer-monomer switch controls CHIP-dependent substrate ubiquitylation and processing.

The high substrate selectivity of the ubiquitin/proteasome system is mediated by a large group of E3 ubiquitin ligases. The ubiquitin ligase CHIP regulates the degradation of chaperone-controlled and chaperone-independent proteins. To understand how CHIP mediates substrate selection and processing, we performed a structure-function analysis of CHIP and addressed its physiological role in Caenorhabditis elegans and human cells. The conserved function of CHIP in chaperone-assisted degradation requires dimer formation to mediate proteotoxic stress resistance and to prevent protein aggregation. The CHIP monomer, however, promotes the turnover of the membrane-bound insulin receptor and longevity. The dimer-monomer transition is regulated by CHIP autoubiquitylation and chaperone binding, which provides a feedback loop that controls CHIP activity in response to cellular stress. Because CHIP also binds other E3 ligases, such as Parkin, the molecular switch mechanism described here could be a general concept for the regulation of substrate selectivity and ubiquitylation by combining different E3s.

A novel curricular framework to develop grant writing skills among MD-PhD students.

Background/Objectives: Physician-scientists have long been in high demand owing to their role as key drivers of biomedical innovation, but their dwindling prevalence in research and medical communities threatens ongoing progress. As the principal avenue for physician-scientist development, combined MD-PhD training programs and NIH-funded Medical Scientist Training Programs (MSTPs) must address all aspects of career development, including grant writing skills. Methods: The NIH F-series grants - the F30 grant in particular - model the NIH format of federal funding, and are thus ideal opportunities to acquire biomedical research grant preparation experience. Therefore, in this report, we describe a curricular model through which predoctoral MSTP students obtain exposure to - and training for - F-series grant conceptualization, writing, and evaluation. Results: Since the development of these longitudinal courses, we observed trending improvements in student funding success rates, particularly among original submissions, and perceived benefits among participating students.

Fostering a diverse regional community of physician-scientist trainees.

Among the many academic challenges faced by dual-degree MD-PhD students is access to professional support networks designed to overcome the unique academic and personal barriers to physician-scientist training. In the current study, we hypothesized that regional access to a student MD-PhD conference, termed the Southeastern Medical Scientist Symposium (SEMSS), would enhance medical and/or graduate training by fostering such relationships between physician-scientist trainees, doing so by discussing both the challenges of physician-scientist training and effective strategies to overcome them. In the current study, we used a mixed-methods approach to evaluate the overall usefulness of SEMSS over a ten-year period (2010-2020) to identify key areas of particular benefit to trainees. The authors used conference registration data to compile self-reported demographic and regional attendance, followed by a post-conference survey to gauge attendee satisfaction. Over the reporting period, SEMSS was attended by equivalent proportions of MD-PhD and undergraduate students, among which were a high-percentage of students from underrepresented minority (URM) groups relative to the national MD-PhD applicant pool; nearly one-third of URM students attendees later matriculated into MD-PhD programs, far exceeding the national MD-PhD matriculation rate. Among the benefits reported by students were "opportunities to network with peers" and opportunities to learn about the physician-scientist career track. Therefore, we therefore propose regional MD-PhD conferences as an effective model to promote diversity within the physician-scientist training pipeline.

LRRK2 dynamics analysis identifies allosteric control of the crosstalk between its catalytic domains.

The 2 major molecular switches in biology, kinases and GTPases, are both contained in the Parkinson disease-related leucine-rich repeat kinase 2 (LRRK2). Using hydrogen-deuterium exchange mass spectrometry (HDX-MS) and molecular dynamics (MD) simulations, we generated a comprehensive dynamic allosteric portrait of the C-terminal domains of LRRK2 (LRRK2RCKW). We identified 2 helices that shield the kinase domain and regulate LRRK2 conformation and function. One helix in COR-B (COR-B Helix) tethers the COR-B domain to the αC helix of the kinase domain and faces its activation loop, while the C-terminal helix (Ct-Helix) extends from the WD40 domain and interacts with both kinase lobes. The Ct-Helix and the N-terminus of the COR-B Helix create a "cap" that regulates the N-lobe of the kinase domain. Our analyses reveal allosteric sites for pharmacological intervention and confirm the kinase domain as the central hub for conformational control.

Drugging the Undruggable: How Isoquinolines and PKA Initiated the Era of Designed Protein Kinase Inhibitor Therapeutics.

In 1984, Japanese researchers led by the biochemist Hiroyoshi Hidaka described the first synthetic protein kinase inhibitors based on an isoquinoline sulfonamide structure (Hidaka et al. Biochemistry, 1984 Oct 9; 23(21): 5036-41. doi: 10.1021/bi00316a032). These led to the first protein kinase inhibitor approved for medical use (fasudil), an inhibitor of the AGC subfamily Rho kinase. With potencies strong enough to compete against endogenous ATP, the isoquinoline compounds established the druggability of the ATP binding site. Crystal structures of their protein kinase complexes, including with cAMP-dependent protein kinase (PKA), showed interactions that, on the one hand, could mimic ATP but, on the other hand, could be optimized for high potency binding, kinase selectivity, and diversification away from adenosine. They also showed the flexibility of the glycine-rich loop, and PKA became a major prototype for crystallographic and nuclear magnetic resonance (NMR) studies of protein kinase mechanism and dynamic activity control. Since fasudil, more than 70 kinase inhibitors have been approved for clinical use, involving efforts that progressively have introduced new paradigms of data-driven drug discovery. Publicly available data alone comprise over 5000 protein kinase crystal structures and hundreds of thousands of binding data. Now, new methods, including artificial intelligence techniques and expansion of protein kinase targeting approaches, together with the expiration of patent protection for optimized inhibitor scaffolds, promise even greater advances in drug discovery. Looking back to the time of the first isoquinoline hinge binders brings the current state-of-the-art into stark contrast. Appropriately for this Perspective article, many of the milestone papers during this time were published in Biochemistry (now ACS Biochemistry).

Early life stress in mice alters gut microbiota independent of maternal microbiota inheritance.

Exposure to early life stress (ELS) is associated with a greater risk of chronic disease development including depression and cardiovascular disease. Altered gut microbiota has been linked to both depression and cardiovascular disease in mice and humans. Rodent models of early life neglect are used to characterize the mechanistic links between early life stress (ELS) and the risk of disease later in life. However, little is understood about ELS exposure and the gut microbiota in the young mice and the influence of the maternal inheritance of the gut microbiota. We used a mouse model of ELS, maternal separation with early weaning (MSEW), and normally reared mice to determine whether the neonate microbiota is altered, and if so, are the differences attributable to changes in dam microbiota that are then transmitted to their offspring. Individual amplicon sequence variants (ASVs) displayed differential abundance in the microbiota of MSEW compared with normally reared pups at postnatal day (PD) 28. Additionally, ELS exposure reduced the alpha diversity and altered microbial community composition at PD28. The composition, levels of alpha diversity, and abundance of individual ASVs in the microbiota of dams were similar from MSEW or normally reared cohorts. Thus, the observed shifts in the abundance of individual bacterial ASVs in the neonates and young pups are likely driven by endogenous effects of MSEW in the offspring host and are not due to inherited differences from the dam. This knowledge suggests that exposure to ELS has a direct effect on microbial factors on the risk of chronic disease development.

Cyclic quantum causal models.

Causal reasoning is essential to science, yet quantum theory challenges it. Quantum correlations violating Bell inequalities defy satisfactory causal explanations within the framework of classical causal models. What is more, a theory encompassing quantum systems and gravity is expected to allow causally nonseparable processes featuring operations in indefinite causal order, defying that events be causally ordered at all. The first challenge has been addressed through the recent development of intrinsically quantum causal models, allowing causal explanations of quantum processes - provided they admit a definite causal order, i.e. have an acyclic causal structure. This work addresses causally nonseparable processes and offers a causal perspective on them through extending quantum causal models to cyclic causal structures. Among other applications of the approach, it is shown that all unitarily extendible bipartite processes are causally separable and that for unitary processes, causal nonseparability and cyclicity of their causal structure are equivalent.

Molecular Basis for Ser/Thr Specificity in PKA Signaling.

cAMP-dependent protein kinase (PKA) is the major receptor of the second messenger cAMP and a prototype for Ser/Thr-specific protein kinases. Although PKA strongly prefers serine over threonine substrates, little is known about the molecular basis of this substrate specificity. We employ classical enzyme kinetics and a surface plasmon resonance (SPR)-based method to analyze each step of the kinase reaction. In the absence of divalent metal ions and nucleotides, PKA binds serine (PKS) and threonine (PKT) substrates, derived from the heat-stable protein kinase inhibitor (PKI), with similar affinities. However, in the presence of metal ions and adenine nucleotides, the Michaelis complex for PKT is unstable. PKA phosphorylates PKT with a higher turnover due to a faster dissociation of the product complex. Thus, threonine substrates are not necessarily poor substrates of PKA. Mutation of the DFG+1 phenylalanine to ß-branched amino acids increases the catalytic efficiency of PKA for a threonine peptide substrate up to 200-fold. The PKA Cα mutant F187V forms a stable Michaelis complex with PKT and shows no preference for serine versus threonine substrates. Disease-associated mutations of the DFG+1 position in other protein kinases underline the importance of substrate specificity for keeping signaling pathways segregated and precisely regulated.

Homoplasmy of the Mitochondrial DNA Mutation m.616T>C Leads to Mitochondrial Tubulointerstitial Kidney Disease and Encephalopathia.

Autosomal-dominant tubulointerstitial kidney disease -(ADTKD) describes tubulointerstitial kidney disease with autosomal-dominant inheritance. In 2017, the term mitochondrial tubulointerstitial kidney disease (MITKD) was introduced for tubulointerstitial kidney disease caused by mitochondrial DNA (mtDNA) mutations. To date, there are few mutations described in literature causing MITKD, one of them is m.616T>C. A 5-year-old girl presented with chronic renal insufficiency and epilepsia. At the age of 3 years, status epileptic occurred and evolved into epilepsia partialis continua. At the age of 5 years, chronic renal failure (CKD II-III) was diagnosed due to tubulointerstitial kidney disease. Urine analysis showed elevated fractional excretions of sodium and chloride. Kidneys were enlarged and hyperechogenic. Blood pressure was elevated. The family history was unremarkable for renal and/or neurological disorders. Genetic testing was performed and revealed homoplasmy of the substitution m.616T>C in our patient's mtDNA. This mutation has been shown to cause chronic tubulointerstitial kidney disease leading to end-stage renal disease (ESRD) and epilepsia formerly. MITKD is a rare mitochondrial disease leading to ESRD and should be suggested in patients with epilepsia and renal insufficiency.

Gamification: An Innovative Approach to Reinforce Clinical Knowledge for MD-PhD Students During Their PhD Research Years.

A longstanding challenge facing MD-PhD students and other dual-degree medical trainees is the loss of clinical knowledge that occurs during the non medical phases of training. Academic medical institutions nationwide have developed continued clinical training and exposure to maintain clinical competence; however, quantitative assessment of their usefulness remains largely unexplored. The current study therefore sought to both implement and optimize an online game platform to support MD-PhD students throughout their research training. Sixty three current MD-PhD students completing the PhD research phase of training were enrolled in an institutionally-developed online game platform for 2 preliminary and 4 competition rounds of 3-4 weeks each. During preliminary game rounds, we found that participation, though initially high, declined precipitously throughout the duration of each round, with 37 students participating to some extent. Daily reminders were implemented in subsequent rounds, which markedly improved player participation. Average participation in competition rounds exceeded 35% (23/63) active participants each round, with trending improvement in scores throughout the duration of PhD training. Both player participation and progress through the research phase of the MD-PhD program correlated positively with game performance and therefore knowledge retention and/or acquisition. Coupled with positive survey-based feedback from participants, our data therefore suggest that gamification is an effective tool for MD-PhD programs to combat loss of clinical knowledge during research training.

Clinical factors and biomarkers predict outcome in patients with immune-mediated thrombotic thrombocytopenic purpura.

Immune-mediated thrombotic thrombocytopenic purpura is characterized by severe thrombocytopenia and microangiopathic hemolytic anemia. It is primarily caused by immunoglobin G type autoantibodies against ADAMTS13, a plasma metalloprotease that cleaves von Willebrand factor. However, reliable markers predictive of patient outcomes are yet to be identified. Seventy-three unique patients with a confirmed diagnosis of immune-mediated thrombotic thrombocytopenic purpura between April 2006 and December 2017 were enrolled from the Univeristy of Alabama at Birmingham Medical Center. Clinical information, laboratory values, and a panel of special biomarkers were collected and/or determined. The results demonstrated that the biomarkers associated with endothelial injury (e.g., von Willebrand factor antigen and collagen-binding activity), acute inflammation (e.g., human neutrophil peptides 1-3 and histone/deoxyribonucleic acid complexes), and activation of the complement alternative pathway (e.g., factors Bb and iC3b) were all significantly increased in patients with acute immune-mediated thrombotic thrombocytopenic purpura compared to those in the healthy controls. Moreover, failure to normalize platelet counts within 7 days or failure to markedly reduce serum lactate dehydrogenase by day 5, low total serum protein or albumin, and high serum troponin levels were also predictive of mortality, as were the prolonged activated partial thromboplastin time, high fibrinogen, and elevated serum lactate dehydrogenase, Bb, and sC5b-9 on admission. These results may help to stratify patients for more intensive management. The findings may also provide a framework for future multicenter studies to identify valuable prognostic markers for immune-mediated thrombotic thrombocytopenic purpura.

Optimization of Laboratory Ordering Practices for Complete Blood Count With Differential.

Objectives: To investigate the utilization of CBC and CBC with differential (CBC w/diff) tests at University of Alabama at Birmingham Hospital, and to determine if a reduction in CBC w/diff tests could be achieved without negatively impacting patient care. Methods: The quantity of testing and distribution of repeated tests before, during, and after an educational intervention were compared. Results: CBC w/diff tests were ordered 10-fold more frequently than CBC tests. The trauma burn intensive care unit ordered the most CBC w/diff tests, with repeat tests done every 4 or 12 hours. The educational intervention reduced the number of CBC w/diff tests ordered and tests repeated every 12 hours. Conclusions: The educational intervention changed the ordering practices of CBC w/diff and CBC tests. This was sustained after the intervention and no negative effects on patient care were noted. Similar interventions may lead to optimization of ordering practices of other laboratory tests.

The Pathology Workforce and Clinical Licensure: The Role of the PhD Clinical Laboratorian in the United States.

There has been a recent recognition of the need to prepare PhD-trained scientists for increasingly diverse careers in academia, industry, and health care. The PhD Data Task Force was formed to better understand the current state of PhD scientists in the clinical laboratory workforce and collect up-to-date information on the training and certification of these laboratorians. In this report, we summarize the findings of the PhD Data Task Force and discuss the relevance of the data collected to the future supply of and demand for PhD clinical laboratory scientists. It is clear that there are multiple career opportunities for PhD scientists in academic medical centers, commercial clinical laboratories, biotechnology and pharmaceutical companies, and the federal government. Certified PhD scientists have and will continue to form an important resource for our technologically advancing field, bringing training in scientific methods, and technologies needed for modern laboratory medicine. The data gathered by the PhD Data Task Force will be of great interest to current and future PhD candidates and graduate PhD scientists as they make decisions regarding future career directions.

Management of chronic myeloid leukemia in the setting of pregnancy: when is leukocytapheresis appropriate? A case report and review of the literature.

BACKGROUND: Chronic myeloid leukemia (CML) is a common hematologic malignancy; however, its occurrence during pregnancy is unusual due to its low prevalence in females of childbearing age. There are conflicting reports of how to best manage CML in pregnancy, particularly in the setting of leukocytosis. HEMAPHERESIS: A 30-year-old female was diagnosed with CML at 18 weeks' estimated gestational age. On initial presentation she reported fatigue, night sweats, and early satiety, and was found to have a white blood cell (WBC) count of 69.3 × 109 /L and platelet count of 366 × 109 /L. Her disease was managed during pregnancy using interferon-α alone despite persistent leukocytosis. CONCLUSION: CML may be effectively managed during pregnancy, even in the setting of leukocytosis, without the application of leukocytapheresis. Management relies not only upon the coordination of drug therapy and fetal monitoring, but requires close communication between multiple medical disciplines. Leukocytapheresis has been safely performed during pregnancy and may be a suitable adjunct management strategy in pregnant patients diagnosed with CML with specific clinical presentations, such as hyperleukocytosis (WBC count > 150 × 109 /L) and/or symptomatic leukostasis.

The Xenobiotic Transporter Mdr1 Enforces T Cell Homeostasis in the Presence of Intestinal Bile Acids.

CD4+ T cells are tightly regulated by microbiota in the intestine, but whether intestinal T cells interface with host-derived metabolites is less clear. Here, we show that CD4+ T effector (Teff) cells upregulated the xenobiotic transporter, Mdr1, in the ileum to maintain homeostasis in the presence of bile acids. Whereas wild-type Teff cells upregulated Mdr1 in the ileum, those lacking Mdr1 displayed mucosal dysfunction and induced Crohn's disease-like ileitis following transfer into Rag1-/- hosts. Mdr1 mitigated oxidative stress and enforced homeostasis in Teff cells exposed to conjugated bile acids (CBAs), a class of liver-derived emulsifying agents that actively circulate through the ileal mucosa. Blocking ileal CBA reabsorption in transferred Rag1-/- mice restored Mdr1-deficient Teff cell homeostasis and attenuated ileitis. Further, a subset of ileal Crohn's disease patients displayed MDR1 loss of function. Together, these results suggest that coordinated interaction between mucosal Teff cells and CBAs in the ileum regulate intestinal immune homeostasis.