Nevin Manimala

FASEB J. 2022 May;36 Suppl 1. doi: 10.1096/fasebj.2022.36.S1.L7744.

ABSTRACT

The lungs are constantly exposed to a diversity of microbes. On average, the human inhales between 0.7 and 7000 bacterial colony forming units (CFU) every minute. The airway epithelium and the airway surface liquid (ASL) which lines the luminal surface, play a vital role in the defence against these inhaled organisms. Glucose concentration in the ASL is much lower than that of blood (approximately 12.5 times lower). It was proposed that low glucose concentration in the ASL contributes to innate protection against the growth of pathogenic organisms which can utilise glucose for growth. Previous research demonstrated that a sustained increase in blood glucose concentration (such as diabetes) led to increased glucose concentration found in the ASL in both human and animals. We therefore hypothesised that the microbial population of the lung would change in the diabetic lung. Seven-week-old female db/db (BKS.Cg-+Leprdb/+Leprdb/OlaHsd) and non-diabetic littermates (BKS.Cg-(Lean)/OlaHsd) db/db mice and non-diabetic littermate controls were purchased from Envigo (UK). Mice were maintained in standard animal housing in a 12h light/dark cycle; water and standard rodent chow available ad libitumand allowed to acclimatise for three weeks before lung microbiome collection. Mice were terminated with an overdose of pentobarbital (0.2ml of 100mg/ml i.p.). Blood was collected for glucose measurement. Bronchoalveolar lavage was performed and 1 mL of solution was used to extract bacterial DNA using QIAamp DNA Microbiome Kit (Qiagen). The V3-V4-region of the 16S rRNA gene was amplified and sequenced using 300 bp paired-end reads on the Illumina MiSeq platform. Bioinformatic analysis was performed using Mothur v1.39.5 as per the MiSeq SOP pipeline. After removing of contaminant sequence reads, downstream statistical analyses were performed using R statistical software. The bacterial diversity in BAL samples was highly variable within and between diabetic and non-diabetic mice. Hyperglycaemia did not affect the a-diversity of the lung microbiome (Inverse Simpson rating). However, hyperglycaemia had a significant effect on the b-diversity of lung microbiome (analysed with AMOVA, p=0.011, n=9) with the microbiome from diabetic mice clustering together. At the genus level, bacteria of genus Staphylococcus were more abundant in the normoglycaemic mice (n=9, p=0.019). The genus Pseudomonas were more abundant in diabetic mice (n=9, p=0.028) and Corynebacterium (n=9, p=0.0018), which are frequently found in the lung microbiome as commensal organisms, were decreased. Taken together, these data indicate that sustained hyperglycaemia modifies the lung microbiome, decreasing the abundance of commensal bacteria and promoting the growth of glucose-utilising bacteria such as Pseudomonas which may include potential pathogenic species such as P. aeruginosa.

PMID:35557398 | DOI:10.1096/fasebj.2022.36.S1.L7744

FASEB J. 2022 May;36 Suppl 1. doi: 10.1096/fasebj.2022.36.S1.L7435.

ABSTRACT

The aim of the study is to investigate the effect of silencing of NILCO by siRNA on the MDR1 gene, which plays a role in chemoresistance. Cells of the human colon cancer lines Caco-2 and HT-29 were cultured at 37˚C in DMEM supplemented with 5% FBS in a humid incubator with 5% CO2. Exponentially growing untreated Caco-2 and HT-29 cells were collected and plated (4 x 10⁵ / well in 2 mL) 24 hours before transfection. Plated cells were treated with either double-stranded siRNA targeting Notch1 mRNA, leptin mRNA, IL-1 β mRNA, control siRNA, or Paclitaxel (PTX) alone, Notch1 siRNA+PTX, Leptin siRNA+PTX, IL-1β siRNA+PTX. The siRNAs of Notch1, IL-1β and Leptin, were applied to the cells at a final concentration of 30 nM, and the cells were transfected with siRNA for 48 hours. After transfection, 100 nM Paclitaxel chemotherapeutic agent for HT-29 and 130 nM for Caco-2 was applied to the cells. Cells were harvested for PCR analysis after treatment with PTX for 24 hours. According to our findings, MDR1 gene expression was significantly increased in the paclitaxel group compared to the control group (p<0.05). Silencing of Notch, leptin and IL-1β decreased MDR1 gene expression compared with the control group, but it was not statistically significant. (p>0.05). However, our results indicated that Notch1 siRNA+PTX, IL-1β siRNA+PTX and Leptin siRNA+PTX downregulated MDR gene expression in the Caco-2 cell line compared with PTX alone treated group (p<0.01). In addition, MDR1 gene expression levels were significantly decreased in the Leptin siRNA+PTX and IL-1β siRNA+PTX groups compared with PTX alone group in the HT-29 cell line, but the decrease in the Notch1 siRNA+PTX group was not statistically significant. (p<0.01, p<0.05, p>0.05). In conclusion, knockdown of NILCO genes is effective in regulating MDR1 expression, which is a hallmark in chemoresistance. The present study was supported by Eskisehir Osmangazi University Scientific Research Projects (BAP) Coordination Unit (grant no. 202011D11).

PMID:35557372 | DOI:10.1096/fasebj.2022.36.S1.L7435

FASEB J. 2022 May;36 Suppl 1. doi: 10.1096/fasebj.2022.36.S1.0R581.

ABSTRACT

INTRODUCTION: Regular physical activity is essential for sedentary older adults (aged 65 years and above) who are at the risk for multiple comorbidities. By performing physical activity, older adults can maintain or improve their physical function and mobility that may help them to be independent for a long time and delay the occurrence of disability in the future.

OBJECTIVE: To assess the effect of the eccentric exercises prior to tai chi and yoga performance in the activities of daily living (ADL), balance and gait, and muscle strength in older adults.

METHODS: Nine sedentary independent older adults aged between 60 and 90 were recruited. The study included the medical pre-protocol assessment, the eccentric (ECC) exercise protocol with and without tai chi and yoga, subjective (Health activities of daily living (ADL) difficulty scale), and objective (Tinetti balance and gait, muscle strength) assessments, and medical post-protocol assessment.

RESULTS: The functional limitations, Tinetti balance and gait score, manual muscle strength testing, and active range of motion (AROM) were improved in the medical post-protocol assessment. Muscle strength showed statistically significant differences on the shoulder abductors (P=0.035), the elbow extensors (P=0.046), the hip abductors (P=0.028) and the hip extensors (P=0.03).

CONCLUSIONS: Our findings show low perceived exercises (ECC) prior to tai chi and yoga training are well suited to the sedentary independent older adults with chronic conditions.

PMID:35557356 | DOI:10.1096/fasebj.2022.36.S1.0R581

FASEB J. 2022 May;36 Suppl 1. doi: 10.1096/fasebj.2022.36.S1.R3123.

ABSTRACT

Cancer stem cells (CSCs) and associated tumor microenvironment (TM) have been proposed to mediate cancer progression, and therapeutic resistance rendering them valuable therapeutic targets in head and neck squamous cell carcinomas (HNSCC). Oral CSCs drive their tumorigenic effect via stromal fibroblasts. Phytochemicals like allantoin are extensively used in wound healing modulating fibroblast activity. The present study aimed to evaluate whether allantoin can modulate CSCs and/or TM and subsequently contribute to sensitization of cancer cells to chemotherapeutic agents. Hep2, OECs, DPSC cell lines were used to test the cytotoxicity of allantoin on cancer, oral epithelial cells, and normal dental pulp stem cells, respectively. The effect of allantoin and/or 5-fluorouracil (5FU) on cell cycle, apoptosis and expression of stem cell and apoptotic markers was tested in vitro. Further validation was performed using in vivo 4-nitroquinoline 1-oxide model of tongue squamous cell carcinoma model with assessment of cytotoxic effect of allantoin by flow cytometry, immunohistochemical staining, western-blot and RT-PCR. Allantoin was safe on both normal oral epithelial cells and dental pulp stem cells. Allantoin showed a statistically significant and dose dependent increase in 5FU apoptotic effect via reduction of cancer stem cell markers CD133 and CD44. Regarding cell cycle arrest, there was no statistically significant difference with addition of allantoin to 5FU. In vivo, allantoin down-regulated SOX2, NANOG, ALDH and OCT4 gene expression in tongue squamous cell carcinoma. Additionally, allantoin reduced the expression IL-6, TGF-Β, PDGF, α- SMA and CD163. Our results proposed that high concentration of allantoin drives its antitumorigenic effect. This was mediated by modulating stromal component of TM via down-regulation of trans-differentiation of cancer associated fibroblasts and tumor associated macrophages. These findings shed the light on potential use of allantoin as cytotoxic drug either alone or combined with other chemotherapeutic drugs.

PMID:35557345 | DOI:10.1096/fasebj.2022.36.S1.R3123

FASEB J. 2022 May;36 Suppl 1. doi: 10.1096/fasebj.2022.36.S1.R4751.

ABSTRACT

At the onset of the COVID-19 pandemic, many traditional in-person courses were converted to synchronous, asynchronous, or hybrid online formats. Instructors of a large undergraduate Basic Human Anatomy course at Indiana University – Bloomington leveraged these changes to convert the lecture portion of the course to a flipped-classroom model with two components: asynchronous online course content delivery, and live in-person interactive learning sessions. The in-person interactive learning sessions introduced students to active learning strategies and techniques (drawing, memory matrices, flow charts, diagrams, etc.), and the students actively participated in these sessions through the use of an online interactive learning platform – TopHat. Here we sought to assess the efficacy and outcomes of the active learning sessions on student performance in the course. Basic Human Anatomy (ANAT-A215) is a 5 credit hour undergraduate (baccalaureate) course that has an enrollment of 480 students with 1 large lecture section and 12 smaller lab sections. In Fall 2021, the instructors modified the lecture component of the course into a flipped classroom format. Lecture content was delivered via asynchronous online lectures, and the F2F class time was listed as optional (but strongly encouraged) for attendance. To facilitate greater student-instructor interaction, one half of the class attended on Mondays and the other half attended on Fridays, and participated in similar interactive sessions. Wednesday F2F interactive lecture sessions were open to the entire class. To encourage attendance and participation in the optional F2F interactive lecture sessions, extra credit points could be earned through participation in TopHat anatomy questions. Points were earned for participation only (.5 points per question). TopHat questions were created by the instructors and consisted of both lower order and higher order questions that tested lecture and lab anatomical content. At the end of each lecture exam block of material, the instructors selected multiple TopHat questions at random for extra credit, with a range of 2-4 extra credit points that could be earned per exam block. We found a statistically significant positive correlation between the number of points accumulated for participating in the F2F active learning lecture session questions and lecture exam scores. Using the number of points accumulated via the TopHat questions as a proxy for lecture attendance, we found that more TopHat points accumulated by students resulted in higher mean lecture exam scores – the greatest benefits occurring if students participated in multiple (>3) F2F lecture sessions per block. Students who regularly attended F2F interactive sessions saw the greatest gains in their lecture exam scores. These findings indicate that active learning strategies could be efficacious in supplementing an asynchronous lecture content delivery format. We shared our findings with the class to encourage them to attend the active learning sessions, but encouraging some student populations to attend these sessions remains a challenge. Future research should be conducted on methods to encourage student involvement and participation in the course.

PMID:35557326 | DOI:10.1096/fasebj.2022.36.S1.R4751

FASEB J. 2022 May;36 Suppl 1. doi: 10.1096/fasebj.2022.36.S1.R6090.

ABSTRACT

Roleplay is a technique that allows students to explore realistic situations by interacting with others in a managed way, in order to develop experiences, skills, and test different strategies in a learning environment. In the teaching of clinical anatomy, the use of this teaching methodology as a tool is scarce. Thus, we implemented the use of this technique as a strategy in the teaching and learning process. The objective of this paper was to report our experience based on the use of the roleplay technique in the teaching and learning process of clinical anatomy. An exploratory, cross-sectional study with a quantitative approach was carried out with 61 students from the anatomy discipline. To carry out the proposed activity, the students were divided into four groups, in which each one planned a clinical anatomy situation. To meet these objectives, Roleplay was structured in three parts: planning the implementation of the roleplay technique, application of roleplay, and debriefing. The results showed that roleplay can be used adequately in the teaching of clinical anatomy. It was observed that 98.1% of students approved the use of the technique in teaching anatomy. It was found that 95.1% of respondents reported that the roleplay achieved the learning goals, while 4.9% reported that the learning goals were not achieved. There was no statistically significant difference between the participants’ ages (p>0.05). It was found that 86.9% of respondents would recommend the use of roleplay as a teaching methodology for the next semester. It was found that roleplay is a playful, versatile, and totally viable and feasible teaching methodology as a tool in the teaching and learning process of clinical anatomy. In addition, this training contributes to discussions and reflections in the teaching-learning process, especially regarding the use of active teaching and learning strategies.

PMID:35557322 | DOI:10.1096/fasebj.2022.36.S1.R6090

FASEB J. 2022 May;36 Suppl 1. doi: 10.1096/fasebj.2022.36.S1.R4775.

ABSTRACT

One key issue in medical education is medical student fear of learning and applying neuroanatomy called neurophobia. In a previous study, we found a negative correlation between neurophobia and neuroanatomy self-efficacy (one’s confidence in their ability to complete neuroanatomy tasks). Our findings suggest increasing neuroanatomy self-efficacy may reduce one’s neurophobia. Additionally, the literature shows self-efficacy improves academic achievement and clinical performance in medical school. It is unknown whether experiences before medical school may later influence medical student neuroanatomy self-efficacy. This study aims to fill this literature gap by isolating which premedical experiences most significantly influence neuroanatomy self-efficacy. We hypothesize medical students with premedical neuroanatomy-specific experience will exhibit greater neuroanatomy self-efficacy than students without this experience. Instrumentation to measure neuroanatomy self-efficacy and categorize premedical experiences was developed and administered to medical students (n=233). Premedical experiences were categorized as: neuroscience (NSC), neuroanatomy (NAC), neuroscience and neuroanatomy (NSAC), cadaveric gross anatomy (GR), or anatomy and physiology (AP). An analysis of variance with a Tukey’s post-hoc test was performed to isolate which premedical experience(s) showed significant differences in neuroanatomy self-efficacy compared to students with no previous experience. All statistical tests were performed at the p=0.05 level. IRB approval was obtained. Data analysis showed indirect premedical neuroanatomy exposure as part of a premedical gross anatomy course (GR) resulted in greater neuroanatomy self-efficacy (p=0.01, Cohen’s d= 0.50) compared to students with no previous experience. Interestingly, enrollment in a premedical molecular neuroscience (NSC) or neuroanatomy (NAC) course individually did not show an increase in neuroanatomy self-efficacy. Only medical students with both premedical NSC and NAC experiences (NSAC) demonstrated higher neuroanatomy self-efficacy than students with no premedical experience (p= 0.001, Cohen’s d= 0.52). Premedical exposure to neuroanatomy as part of a gross anatomy course (GR) or enrollment in both premedical neuroanatomy andneuroscience courses (NSAC) appears to be associated with higher neuroanatomy self-efficacy in medical students. Considering the decline in neurologists, and fewer medical students choosing neurology, these findings support youth STEM initiatives to provide increased access to both neuroscience and neuroanatomy experiences along with cadaveric gross anatomy.

PMID:35557316 | DOI:10.1096/fasebj.2022.36.S1.R4775

FASEB J. 2022 May;36 Suppl 1. doi: 10.1096/fasebj.2022.36.S1.0R412.

ABSTRACT

The Fishbowl is an active teaching methodology whose group discussion format promotes dialogue and exchange of experience among session students and allows everyone to have an equal chance to express their opinions and views during class. So, we use this method to teach endocrine system anatomy to undergraduate students. However, there is little evidence of the use of this methodology to guide teachers on the best way to implement the Fishbowl in teaching anatomy. Based on these premises, the objective of this work was to report our experience based on the use of the Fishbowl technique in the teaching and learning process of the anatomy of the endocrine system. A quasi-experimental study with a quantitative approach was carried out with students from the discipline of anatomy of the endocrine system. The technique used was the open fishbowl in online mode. For data analysis, we used the relative frequency and the Mann Whitney test with the aid of the Graph Pad Prism version 7.0 program, with the data with a value of p<0.05 being considered significant. It was observed that there was no statistically significant difference between the participants’ ages (p>0.05). In our studies, we found that Fishbowl was more effective for teaching undergraduate anatomy than traditional teaching (p<0.05). The Fishbowl proved to be a satisfactory teaching technique for students when compared to the traditional class. We found that students who participated in the Fishbowl got statistically significantly more questions on the test when compared to students in traditional education (p<0.05). It was found that the Fishbowl allowed the educational process to be carried out with greater interaction in a more playful way. In addition, students became protagonists in the teaching and learning process, participating more effectively in classes. Thus, the Fishbowl becomes another active tool in the process of teaching and learning anatomy.

PMID:35557314 | DOI:10.1096/fasebj.2022.36.S1.0R412

FASEB J. 2022 May;36 Suppl 1. doi: 10.1096/fasebj.2022.36.S1.R4007.

ABSTRACT

BACKGROUND: Since the seventeenth century, the primary approach to teaching anatomy has involved hands-on learning using cadaveric specimens. However, the ability to use this long-standing tradition was curtailed in the 2020-2021 school year due to the COVID-19 pandemic. Many institutions closed physical classrooms entirely, launching experiential courses, such as anatomy, into the online space.

HYPOTHESIS: We hypothesized that Q-methodology could be used to uncover student perceptions of an introductory anatomy and physiology course that was offered online for the very first time.

METHODS: Q-methodology, considered the study of subjectivity, is an approach that statistically uncovers groups of individuals with shared perceptions within a larger cohort. Instructors can use Q-methodology to identify groups of students with shared needs, allowing for more specific and productive course reform. In the current study, Q-methodology was used as a means of course evaluation in the fall 2020 and winter 2021 semesters. Students were asked to sort 44 opinion-based statements in a quasi-normal table based on their level of agreement. By-person factor analysis of 166 responses revealed three statistically distinct groups of students.

RESULTS: The three groups were assigned the following monikers: Connected and Contented (CC), Disconnected and Disgruntled (DD), and Interconnected and Collaborative (IC). CC students (n=66) felt generally ambivalent toward course components and were comfortable with the technology skills required to participate in the online course space. DD students (n=50) were deeply unhappy with several elements of the course, including lectures, assignments, and evaluations. These students also felt as if they were teaching themselves. Finally, IC students (n=29) looked favourably upon the tutorial space and the role of teaching assistants. Analysis also revealed that some sentiments were shared across all three groups, including the preference for physical rather than virtual specimens, and the desire for more practice questions from faculty in order to prepare for bellringer exams. Interestingly, cohort opinions did not remain static across both semesters. There was a positive attitude shift as more students felt “Disconnected and Disgruntled” in the fall, and “Connected and Contented” in the winter.

CONCLUSIONS: These findings are useful for anatomy instructors interested in transitioning courses to an online or blended space, particularly in the face of ever evolving public health restrictions. The current study also models the wealth of information that can be uncovered using Q-methodology – useful for anyone interested in the previously amorphous study of subjectivity.

PMID:35557298 | DOI:10.1096/fasebj.2022.36.S1.R4007

FASEB J. 2022 May;36 Suppl 1. doi: 10.1096/fasebj.2022.36.S1.R4298.

ABSTRACT

SAMP1/YitFC (SAMP) mice exhibit spontaneous ileitis as they age with morphological and functional epithelial abnormalities mimicking human Crohn’s disease, a chronic disease characterized by ileal inflammation. Indole-3-carbinole (I3C), found in broccoli and Brussel sprouts, plays a crucial role in the anti-inflammatory processes by activating the aryl hydrocarbon receptor (AhR), a transcription factor vital for epithelial and immune cell functions. The impact of I3C diet on disease progression and epithelial integrity in SAMP model of ileitis is not known. METHODS: 6-week-old male and female (n=8 per group) SAMP and control AKR/J mice (Jackson) were fed for 14 weeks either a standard chow diet or a purified diet supplemented with 200 ppm with indole-3-carbinole (+I3C diet), or purified diet depleted of AhR ligands (-I3C diet). Ileum samples were collected for qPCR and histological analyses. Statistical analysis was performed by Tukey’s multiple comparison test for all three diets. To detect changes between +I3C vs. -I3C, delta values (Δ +/-I3C = SAMP-AKR) were analyzed by unpaired T-test. RESULTS: As expected, H&E staining for the ileum of SAMP mice fed chow diet, showed thickening of the muscle layer, neutrophil infiltration, and epithelial tufting vs. AKR mice. Severity of inflammation in SAMP mice, as assessed by fecal calprotectin ELISA, mRNA expression of cytokines, and histological analysis was not significantly impacted by dietary depletion or supplementation of I3C. With respect to epithelial abnormalities, Periodic Acid Schiff staining showed increased number of goblet cells per villi (~47-75%) in SAMP mice on all three diets. While mRNA levels of the major goblet cell secretory proteins, mucin2 (Muc2) and intestinal trefoil factor (ITF), were dramatically decreased in SAMP mice vs. AKR, supplementation of I3C attenuated this decrease in Muc2 and ITF by ~35% and 51% (p<.001), respectively, indicating defective synthesis or secretion. Similarly, I3C diet significantly (p<.01) attenuated the decrease in Paneth cell markers lysozyme C-1 (LYZ1) alpha defensin 5 (Def5α) and regenerating islet-derived protein 3 alpha (Reg3α) in SAMP mice by ~65%, 60%, and 24%, respectively. A decrease in fibroblast growth factor 15 (FGF15) and the epithelial ion transporters serotonin transporter (SERT) and organic solute transporter (OST) were observed in ileal mucosa of SAMP mice vs. AKR, and the supplementation of I3C mitigated these decreases by 30-58% (p<.01). There were no significant changes in the expression of tryptophan hydroxylase 1 (tph-1), C-X-C motif chemokine ligand 2 (CXCL2), or AhR by I3C diet in SAMP mice. In summary, I3C supplementation significantly attenuated the goblet cell, Paneth cell, and epithelial-specific abnormalities seen in SAMP mice. These data imply that dietary supplementation of AhR ligands may be a viable therapeutic strategy for Crohn’s patients to restore epithelial integrity.

PMID:35557290 | DOI:10.1096/fasebj.2022.36.S1.R4298