Improving Student Experiences During USMLE Step 1 "Dedicated Preparation Period" via a Course Designed to Holistically Support Academic and Wellness Needs.

Common medical school practice includes an independent "dedicated preparation period" for the USMLE Step 1 exam during which students have reported considerable stress and anxiety. Carle Illinois College of Medicine sought to improve their students' experience during the dedicated preparation period via a unique course, Phase 1 Synthesis and Summary, designed to provide support for academics and wellness. The strengths of the course were elements that maximized student flexibility, autonomy, wellness, and targeted self-study. The course design reported here may provide a model for other schools to design courses to holistically improve students' preparation for Step 1 and other board exams. Supplementary Information: The online version contains supplementary material available at 10.1007/s40670-023-01791-2.

Potential for mobile health (mHealth) prevention of cardiovascular diseases in Kerala: A population-based survey.

BACKGROUND: India's southern state of Kerala stands at the forefront of India's epidemic of cardiovascular disease (CVD), among other non-communicable diseases (NCDs). Mobile phone use in healthcare (mHealth) has shown promise in India, including NCDs. However, suitability and acceptability of m-Health interventions is poorly researched, particularly in rural settings. OBJECTIVES: METHODS: A questionnaire regarding mobile phone usage and possible use in healthcare was verbally administered in five primary health centres and by home visits in five village councils ("panchayats") of Ernakulam, Kerala. Adults who spoke Malayalam or English, with access to a mobile phone were recruited by convenience sampling in partnership with accredited social health activists (ASHAs). Quantitative data analysis was conducted using SPSS software. RESULTS: 262 participants were recruited. 87% routinely used and 88% owned a mobile phone. 92% were willing to receive mHealth advice, and 94% favoured mobile medication reminders. 70.3% and 73% preferred voice calls over short messaging service (SMS) for delivering health information and medication reminders, respectively. 85.9% would send home recorded information on their blood pressure, weight, medication use and lifestyle to a doctor or ASHA. 75.2% trusted the confidentiality of mHealth data, while 77.1% had no concerns about the privacy of their information. CONCLUSIONS: The majority of this population approve mHealth interventions. While further investigation of mHealth as a health education tool is warranted, SMS interventions may fail to maximise equity and penetration across all patient groups.

A Comparative Study of G-Quadruplex Unfolding and DNA Reeling Activities of Human RECQ5 Helicase.

RECQ5 is one of five members of the RecQ family of helicases in humans, which include RECQ1, Bloom (BLM), Werner (WRN), RECQ4, and RECQ5. Both WRN and BLM have been shown to resolve G-quadruplex (GQ) structures. Deficiencies in unfolding GQ are known to result in DNA breaks and genomic instability, which are prominent in Werner and Bloom syndromes. RECQ5 is significant in suppressing sister chromatid exchanges during homologous recombination but its GQ unfolding activity are not known. We performed single-molecule studies under different salt (50-150 mM KCl or NaCl) and ATP concentrations on different GQ constructs including human telomeric GQ (with different overhangs and polarities) and GQ formed by thrombin-binding aptamer to investigate this activity. These studies demonstrated a RECQ5-mediated GQ unfolding activity that was an order of magnitude weaker than BLM and WRN. On the other hand, BLM and RECQ5 demonstrated similar single-stranded DNA (ssDNA) reeling activities that were not coupled to GQ unfolding. These results demonstrate overlap in function between these RecQ helicases; however, the relatively weak GQ destabilization activity of RECQ5 compared to BLM and WRN suggests that RECQ5 is not primarily associated with GQ destabilization, but could substitute for the more efficient helicases under conditions where their activity is compromised. In addition, these results implicate a more general role for helicase-promoted ssDNA reeling activity such as removal of proteins at the replication fork, whereas the association of ssDNA reeling with GQ destabilization is more helicase-specific.

Potential for the use of mHealth in the management of cardiovascular disease in Kerala: a qualitative study.

OBJECTIVES: To assess the potential for using mHealth in cardiovascular disease (CVD) management in Kerala by exploring: (1) experiences and challenges of current CVD management; (2) current mobile phone use; (3) expectations of and barriers to mobile phone use in CVD management. DESIGN: Qualitative, semistructured, individual interviews. SETTING: 5 primary health centres in Ernakulam district, Kerala, India. PARTICIPANTS: 15 participants in total from 3 stakeholder groups: 5 patients with CVD and/or its risk factors, 5 physicians treating CVD and 5 Accredited Social Health Activists (ASHAs). Patients were sampled for maximum variation on the basis of age, sex, CVD diagnoses and risk factors. All participants had access to a mobile phone. RESULTS: The main themes identified relating to the current challenges of CVD were poor patient disease knowledge, difficulties in implementing primary prevention and poor patient lifestyles. Participants noted phone calls as the main function of current mobile phone use. The expectations of mHealth use are to: improve accessibility to healthcare knowledge; provide reminders of appointments, medication and lifestyle changes; save time, money and travel; and improve ASHA job efficacy. All perceived barriers to mHealth were noted within physician interviews. These included fears of mobile phones negatively affecting physicians' roles, the usability of mobile phones, radiation and the need for physical consultations. CONCLUSIONS: There are three main potential uses of mHealth in this population: (1) as an educational tool, to improve health education and lifestyle behaviours; (2) to optimise the use of limited resources, by overcoming geographical barriers and financial constraints; (3) to improve use of healthcare, by providing appointment and treatment reminders in order to improve disease prevention and management. Successful mHealth design, which takes barriers into account, may complement current practice and optimise use of limited resources.

ATP-dependent G-quadruplex unfolding by Bloom helicase exhibits low processivity.

Various helicases and single stranded DNA (ssDNA) binding proteins unfold G-quadruplex (GQ) structures. However, the underlying mechanisms of this activity have only recently come to focus. We report kinetic studies on Bloom (BLM) helicase and human telomeric GQ interactions using single-molecule Förster resonance energy transfer (smFRET). Using partial duplex DNA (pdDNA) constructs with different 5' ssDNA overhangs, we show that BLM localizes in the vicinity of ssDNA/double-stranded DNA (dsDNA) junction and reels in the ssDNA overhang in an ATP-dependent manner. A comparison of DNA constructs with or without GQ in the overhang shows that GQ unfolding is achieved in 50-70% of reeling attempts under physiological salt and pH conditions. The unsuccessful attempts often result in dissociation of BLM from DNA which slows down the overall BLM activity. BLM-mediated GQ unfolding is typically followed by refolding of the GQ, a pattern that is repeated several times before BLM dissociates from DNA. BLM is significantly less processive compared to the highly efficient GQ destabilizer Pif1 that can repeat GQ unfolding activity hundreds of times before dissociating from DNA. Despite the variations in processivity, our studies point to possible common patterns used by different helicases in minimizing the duration of stable GQ formation.

Asymmetric unwrapping of nucleosomes under tension directed by DNA local flexibility.

Dynamics of the nucleosome and exposure of nucleosomal DNA play key roles in many nuclear processes, but local dynamics of the nucleosome and its modulation by DNA sequence are poorly understood. Using single-molecule assays, we observed that the nucleosome can unwrap asymmetrically and directionally under force. The relative DNA flexibility of the inner quarters of nucleosomal DNA controls the unwrapping direction such that the nucleosome unwraps from the stiffer side. If the DNA flexibility is similar on two sides, it stochastically unwraps from either side. The two ends of the nucleosome are orchestrated such that the opening of one end helps to stabilize the other end, providing a mechanism to amplify even small differences in flexibility to a large asymmetry in nucleosome stability. Our discovery of DNA flexibility as a critical factor for nucleosome dynamics and mechanical stability suggests a novel mechanism of gene regulation by DNA sequence and modifications.

RecQ-core of BLM unfolds telomeric G-quadruplex in the absence of ATP.

Various helicases and single-stranded DNA (ssDNA) binding proteins are known to destabilize G-quadruplex (GQ) structures, which otherwise result in genomic instability. Bulk biochemical studies have shown that Bloom helicase (BLM) unfolds both intermolecular and intramolecular GQ in the presence of ATP. Using single molecule FRET, we show that binding of RecQ-core of BLM (will be referred to as BLM) to ssDNA in the vicinity of an intramolecular GQ leads to destabilization and unfolding of the GQ in the absence of ATP. We show that the efficiency of BLM-mediated GQ unfolding correlates with the binding stability of BLM to ssDNA overhang, as modulated by the nucleotide state, ionic conditions, overhang length and overhang directionality. In particular, we observed enhanced GQ unfolding by BLM in the presence of non-hydrolysable ATP analogs, which has implications for the underlying mechanism. We also show that increasing GQ stability, via shorter loops or higher ionic strength, reduces BLM-mediated GQ unfolding. Finally, we show that while WRN has similar activity as BLM, RecQ and RECQ5 helicases do not unfold GQ in the absence of ATP at physiological ionic strength. In summary, our study points to a novel and potentially very common mechanism of GQ destabilization mediated by proteins binding to the vicinity of these structures.

Insight into helicase mechanism and function revealed through single-molecule approaches.

Helicases are a class of nucleic acid (NA) motors that catalyze NTP-dependent unwinding of NA duplexes into single strands, a reaction essential to all areas of NA metabolism. In the last decade, single-molecule (sm) technology has proven to be highly useful in revealing mechanistic insight into helicase activity that is not always detectable via ensemble assays. A combination of methods based on fluorescence, optical and magnetic tweezers, and flow-induced DNA stretching has enabled the study of helicase conformational dynamics, force generation, step size, pausing, reversal and repetitive behaviors during translocation and unwinding by helicases working alone and as part of multiprotein complexes. The contributions of these sm investigations to our understanding of helicase mechanism and function will be discussed.

BLM helicase measures DNA unwound before switching strands and hRPA promotes unwinding reinitiation.

Bloom syndrome (BS) is a rare genetic disorder characterized by genomic instability and a high predisposition to cancer. The gene defective in BS, BLM, encodes a member of the RecQ family of 3'-5' DNA helicases, and is proposed to function in recombinational repair during DNA replication. Here, we have utilized single-molecule fluorescence resonance energy transfer microscopy to examine the behaviour of BLM on forked DNA substrates. Strikingly, BLM unwound individual DNA molecules in a repetitive manner, unwinding a short length of duplex DNA followed by rapid reannealing and reinitiation of unwinding in several successions. Our results show that a monomeric BLM can 'measure' how many base pairs it has unwound, and once it has unwound a critical length, it reverses the unwinding reaction through strand switching and translocating on the opposing strand. Repetitive unwinding persisted even in the presence of hRPA, and interaction between wild-type BLM and hRPA was necessary for unwinding reinitiation on hRPA-coated DNA. The reported activities may facilitate BLM processing of stalled replication forks and illegitimately formed recombination intermediates.

Mapping nucleosome locations on the 208-12 by AFM provides clear evidence for cooperativity in array occupation.

Concatameric sea urchin 5S rDNA templates reconstituted with histones provide very popular chromatin models for many kinds of in vitro studies. We have used AFM to characterize the locational aspects of nucleosome occupation on one such array, the 208-12, by determining the internucleosomal- and end-distance distributions for arrays reconstituted to various subsaturating levels with nonacetylated or hyperacetylated HeLa histones. A simulation analysis of the experimental distributions confirms the qualitative conclusions and provides quantitative parameter values for the identified features. For nonacetylated arrays, the end-distance data demonstrate the nucleosome positioning ability of the 5S sequence and detect an enhanced preference for nucleosomes to bind at DNA termini. The internucleosomal-distance data provide clear evidence for cooperativity in nucleosome location on these templates, detectable even at subsaturated loading levels. Hyperacetylated arrays show no change in the preference of nucleosomes to bind at termini and a slight change in nucleosome positioning behavior but, most strikingly, little or no evidence for cooperativity in nucleosome location. Thus, acetylation of the N-terminal histone tails abolishes the cooperativity.

Radionuclide studies in the evaluation of urinary tract infections.

OBJECTIVE: To estimate the prevalence of vesicoureteral reflux (VUR) and renal scarring in children presenting with culture proven urinary tract infections (UTI). DESIGN: Descriptive study. SETTING: Tertiary care hospital-based study. SUBJECTS: Thirty-two children with proven UTI were evaluated by means of an abdominal ultrasonogram (USG), Technetium-99m Dimercapto Succinic Acid (DMSA) scan and Direct Radionuclide Cystography (DRCG). A micturating cystourethrogram (MCU) was performed to rule out any structural abnormality and to grade VUR. RESULTS: A total of 64 renal units in 32 children were evaluated. DMSA scan showed scarring in 27 renal units (42.2%) in 16 patients. Bilateral renal scarring was more common in older (> 2 yr) children as compared to younger ones (89% Vs 43%; p < 0.05). USG detected abnormalities in 13 renal units (20.3%) in 7 cases. VUR was detected in 37.5% of children of all age group by DRCG. In contrast, MCU showed evidence of VUR in only 13/20 renal units with a sensitivity of 65% as compared to DRCG and did not pick up any additional VUR that could have been missed on the DRCG. Only 3/9 in < 2 yr, in contrast to 10/11 in > 2 yr were positive for VUR on MCU (p < 0.05). However, MCU detected evidence of cystitis in 3 children and a bladder diverticulum in one patient. CONCLUSION: Wherever available, DMSA scan should be considered as a part of the first line investigations in any patient presenting with UTI. DRCG can also be performed in the same sitting to screen for the presence of reflux particularly for girls.