bluuebunny/crossref_metadata_2025 huggingface dataset:
Glucose homeostasis is impaired in Guanylyl Cyclase/Natriuretic Peptide Receptor‐A Gene‐Knockout and Gene‐Duplication Mutant Mice
Authors: Ramasamy Chandramohan , Neelamegam Kandasamy , Bloodworth Meaghan , Pandey Kailash N.
Published: May 2022

Atrial and brain natriuretic peptides (ANP and BNP) are produced in the heart, regulate blood pressure (BP) and fluid homeostasis through vasodilatory and diuretic actions. Both ANP and BNP act by binding to guanylyl cyclase/natriuretic peptide receptor‐A (GC‐A/NPR‐A). Systemic disruption of Npr1(encoding GC‐A/NPRA) leads to volume overload, high BP, and congestive heart failure. However, the underlying mechanisms are not yet precisely determined. The aim of this study was to investigate whether Npr1plays a critical role in regulating glucose homeostasis in Npr1 gene‐disrupted mice. The adult male and female (16‐18 wk) Npr1 gene‐knockout haplotype (Npr1+/‐, 1‐copy), wild‐type (Npr1+/+, 2‐copy), and gene‐duplicated (Npr1+ +/+ +, 4‐copy) mice were fasted for 16 h and given free access to water. The glucose administration was done both orally and intraperitoneally (IP: 2 g/kg body weight) in mice to determine oral glucose tolerance test (OGTT) and IP glucose tolerance test (IPGTT). The glucose levels in blood were determined by performing tail bleeds at 0, 15, 30, 60, 90, and 120 min using the AlphaTRAK blood glucose monitoring system (Zoetis Inc, Kalamazoo, MI). Systolic BP (SBP) was determined by non‐invasive computerized tail‐cuff method (Visitech 2000). The results showed that the blood glucose level was elevated to the maximum value at 15 min after glucose administration. Glucose levels were declined to near base line at 120 min in 2 copy mice (OGTT: 102 ± 3 mg/dL in male and 97 ± 4 mg/dL in female, IPGT: 101 ± 4 mg/dL in male and 96 ± 3 mg/dL in female). However, in 1‐copy mice, the blood glucose levels remained elevated even after 120 min (OGTT: 248 ± 5 mg/dL in male and 226 ± 3 mg/dL in female, IPGT: 254 ± 4 mg/dL in male and 229 ± 5 mg/dL in female) compared with 2‐copy mice. The blood glucose levels were also significantly lower at 120 min in 4‐copy mice (OGTT: 77 ± 4 mg/dL in male and 74 ± 3 mg/dL in female, IPGT: 75 ± 3 mg/dL in male and 71 ± 2 mg/dL in female) compared with 2‐copy mice. SBP was significantly greater in 1‐copy mice (137 + 4 mmHg in male and 126 + 3 mmHg in female) than 2‐copy mice (100 + 2 mmHg in male and 91 + 2 mmHg in female). SBP was significantly lower in 4‐copy mice (83 + 2 mmHg in male and 77 + 2 mmHg in female) than 2‐copy mice. OGTT showed a significantly lower level of maximal blood glucose compared with IPGTT. The findings of the present study suggest that Npr1 plays a critical role in the regulation of glucose homeostasis and the loss of Npr1 significantly increases the glucose levels in mutant animals.
Role of EnvZ/OmpR Two‐component System in Antibiotic Resistance in Salmonella enterica Serovar Enteritidis
Authors: Ko Duhyun , Choi Garam , Sung Dayoung , Choi Sang Ho
Published: May 2022

Outer membrane porins (OMPs) form water‐filled open channels and allow the entry of small hydrophilic antibiotics. Thus, compositional modulation of OMPs is frequently observed in multidrug‐resistant bacteria, including a human pathogen Salmonella enterica. However, the mechanism by which S. entericamodulates OMP composition to enhance antibiotic resistance has not been well elucidated. In this study, we found that the S. enterica serovar Enteritidis strain with an active state of EnvZ/OmpR is highly resistant to various antibiotics, especially β‐lactams. Transcriptome analysis discovered that the active state of EnvZ/OmpR induces differential expression of multiple OMP genes, including SEN1522, SEN2875, ompD, and ompW. Biochemical analyses identified that OmpR activates SEN1522 and SEN2875 but represses ompD and ompW by directly binding to their upstream regions. Interestingly, the cellular level of OmpR determines the expression of the four OMP genes, and particularly, the phosphorylation of OmpR is not required for the repression of ompD and ompW. Phenotypic analysis revealed that a decreased expression of ompD is responsible for the EnvZ/OmpR‐mediated resistance to β‐lactams in S. Enteritidis. Notably, EnvZ/OmpR represses the expression of ompD upon exposure to β‐lactams and presumably provides S. Enteritidis with benefits for survival. Altogether, this study suggests that EnvZ/OmpR remodels OMP composition in response to β‐lactams and thereby enhances resistance in S. Enteritidis to the antibiotics.
Syndecan‐4 functionalized elastomeric biomaterial promotes in situ endovascular tissue repair and regeneration
Authors: Wu Yidi , Wagner William
Published: May 2022

While many acellular endovascular biomaterials have been developed to attract endothelial precursors cells (EPCs) to promote tissue repair and regeneration, none of them truly create a conducive microenvironment for EPC adhesion, growth, and differentiation. Syndecan‐4 is a heparan sulfate containing proteoglycan with high affinity and binding specificity for various growth factors, cytokines, chemokines, is a significant component of the endothelial cell glycocalyx. In this study, an endovascular biomaterial composite of type I collagen, poly (glycerol sebacate) and silk fibroin, termed PFC, was functionalized with syndecan‐4 and stromal cell‐derived factor 1 alpha (SDF‐1α) to study the interaction with EPCs. The functionalized biomaterial (PFC_SYN4) significantly promoted the adhesion of EPCs. PFC_SYN4 captured 50% more cells than PFC under dynamic binding conditions (P<0.001) and 20% more cells than PFC under physiological flow conditions (P<0.05). Competitive binding studies showed EPCs bound to PFC_SYN4 in the presence of excess leucocytes. After 2 hours of incubation, average cell spreading area of EPCs on PFC_SYN4 (1252 ± 415 µm2) was significantly greater (P<0.05) than on PFC (1149 ± 392 µm2). At day 4, vWF expression was doubled for cells on PFC and PFC_SYN4. CD31 was upregulated about 3.4 fold in cells cultured on PFC and 7.2 fold in cells cultured on PFC_SYN4 (P<0.05), at day 7, CD31 and vWF expression was significantly greater than on day 4. PFC_SYN4 increased 10.6 fold for vWF and PFC increased 4 fold. PFC_SYN4 had doubled amount of CD31 expression compared to PFC (P<0.05). This study expands our knowledge of the potential benefit of using syndecan‐4 to functionalize acellular biomaterials and facilitate the adhesion, growth, and differentiation of precursor cells in order to further promote tissue repair and regeneration.
Head and Neck Cancer‐derived Small Extracellular Vesicles Activate TRPV1+ neurons to Mediate Cancer Pain
Authors: Laumet Geoffroy , Inyang Kufreobong E. , Evans Christine M. , Heussner Matthew , Petroff Margaret , Reimers Mark , Tykocki Nathan , Vermeer Paola , Folger Joseph K.
Published: May 2022

Most patients with head and neck squamous cell carcinoma (HNSCC) experience pain and pain is associated with a poor prognosis. Despite the frequency and severity of HNSCC pain, current treatments fail to adequately control pain. Cancer‐derived small extracellular vesicles (sEVs, size 30‐150 nm) are well‐positioned to be a mediator of communication between cancer cells and neurons. We hypothesize that cancer‐derived sEVs contribute to cancer pain. Mouse oropharyngeal cells were retrovirally transduced to stably express HPV16 viral oncogenes, E6 and E7, H‐Ras and luciferase and therefore called mEERL cells. Implantation of mEERL cells into WT mice induces evoked and spontaneous pain. Administration of the sEV release inhibitor GW4869 attenuates the pain in tumor‐bearing mice. Additionally, blocking sEV release specifically in the cancer cells, by deleting Rab27a and Rab27b, two proteins required for exosome release, significantly delayed the development of pain hypersensitivity. To test whether cancer‐derived sEVs are sufficient to induce pain, we isolated sEVs from mEERL culture and injected them subcutaneously. Injection of isolated sEVs triggers pain hypersensitivity in both sexes. NSAID ketoprofen has no effect on sEVs‐induced pain hypersensitivity. In contrast blocking nociceptor neuron activity with the membrane‐impermeant lidocaine derivate QX‐314 alleviates sEVs‐induced pain hypersensitivity. In primary culture of trigeminal ganglion neurons, addition of cancer‐derived sEVs induces expression of the neuronal injury marker activating transcription factor 3 (ATF3) and calcium influx measured by calcium imaging in Trpv1Cre:GCaMP6 mice. Given that sEVs activate TRPV1+ neurons (mostly nociceptors), we examine the impact of TRP1V1+ neuron ablation on cancer pain. Chemical ablation of TRPV1+ neurons by resiniferatoxin (RTX) treatment prevents the development of evoked and spontaneous pain in tumor‐bearing mice. Finally, to further explore the potential mechanism of nociception triggered by sEVs, we used published human RNA‐sequencing data to investigate the change in gene expression in human cultured sensory neurons exposed to sEVs. Ingenuity pathways analysis (IPA) identified several pathways linked to the initiation of translation, a pathway known to contribute to nociception and neuroplasticity associated with chronic pain. Pharmacological inhibition of translation by rapamycin (mTOR inhibitor), and narciclasine (AMPK activator), alleviates and prevents pain in tumor‐bearing mice respectively. In summary, our study shows that cancer‐derived sEVs directly activate TRPV1+ neurons to trigger cancer pain and identify new actionable pharmacological targets.
Structural and functional analysis of cancer related proteins: A critical target for drug development
Authors: Sharma Ruby , Saxena Ajay K.
Published: May 2022

Protein kinases are involved in cancer metabolism and have been targeted for structure‐based drug development. The crystals structures of several clinically important protein kinase‐inhibitor complexes have been determined, which provide the mechanism of inhibition of protein kinases. In our laboratory, we are working on structure and function analysis on two protein kinases e.g. (i) a non‐receptor tyrosine kinase expressed in hepatocellular carcinoma (TEC) and Class I Phosphoinositide 3‐Kinase (PI3K). We have cloned and expressed both kinases in E. coli and looking forward for biochemical and structural characterization of both proteins.

In addition, we are also working on structural and biochemical characterization of Ergp55 protein belongs to Ets family of transcription factor. The Ergp55 is involved in various development processes and regulation of cancer metabolism. Using biochemical assay, we have identified the part of Ergp55 protein that mediates the auto‐inhibition using E74 and cfos promoter DNA sequences. We have also crystallized and solved the crystal structure of ETS domain of Ergp55 in native and its complexes with E74 and cFOS promoter DNA sequences. The structure analysis provides the insights into DNA sequence discrimination by ETS domain of Ergp55 protein. These studies will aid in designing the compounds, which stabilize the inhibited form of Ergp55 and inhibits its binding to DNA. Based on crystal structures, we have designed the DNA binding compounds, which interfere in ETS‐DNA binding and proved to be potential drugs against prostate cancer.
Activity Monitoring During the Instruction of Gross Anatomy
Authors: Yaggie James A.
Published: May 2022

Over the past decades, a compendium of various physical activities has been compiled that ranges from more vigorous activities in the occupational and recreational domains, to mundane household tasks and chores. These have been developed to allow users to understand their individual activity profile for intake and caloric expenditure. Teaching gross anatomical dissection courses is physically demanding, and anecdotally, has a different activity profile than teaching other courses. The purpose of this study was to quantify the basic activity profile of gross anatomical instruction and compare these measure to other occupation tasks. For a period of 4‐years, faculty and laboratory staff (n=52; 35.6 years + 8) were provided commercially available activity monitors (Fitbit Charge, Fitbit, San Francisco, CA) and were worn during all gross anatomy lab dissections (272 min/session + 38) in a semester‐long class. Data were collected and reported after each class session including, steps, caloric expenditure and time of activity. The average caloric expenditure/session was 569.47 kCal (SD = 33.8), or 126.55 kCal/hour during the instruction of gross anatomy lab experiences and was found to be similar to general carpentry (145 kCal/hour) and custodial work (136 kCal/hour). However, the profession that was most similar was nursing. A recent study has shown (Allan, et al, Journal of Research in Nursing, 2019) that ward nurses on 12‐hour shifts exhibit caloric expenditure of 1512kCal/shift, or 126.75 kCal/hour. When the objective tasks are compared, its not surprising that these professions exhibit similar results. Both repeatedly perform work from station‐to‐station throughout the day requiring above average amounts of standing and walking. Further, the work performed at a station also requires a mixture of finite precision and moving of the subject/patient. Although these data aren’t unique or novel, it may be of particular interest to those that perform this occupation skill. Guiding the dissection experiences for medical and allied health students is a vigorous and strenuous job that requires the background and familiarity human anatomical structures with physical demands that are more extensive than a typical class or laboratory setting.
A 24‐Hour Fast Increases Liver Cytotoxic T Cell Populations in Genetically Obese Mice: A Preliminary Study
Authors: Graham Elliot L. , Trikha S. Raj J. , Risk Briana , Weir Tiffany L. , Gentile Christopher L.
Published: May 2022

BACKGROUND

Recent data suggests that regulated fasting protocols can improve metabolic parameters associated with obesity. However, questions remain regarding the appropriate fasting duration needed to attenuate obesity‐associated complications such as glucose intolerance, arterial stiffness, and immune cell dysfunction. Therefore, the primary purpose of this study was to examine if an acute 24‐hour fast improves detrimental consequences of obesity and alters tissue‐specific immune cell counts in obese mice.

METHODS

Ten 12‐week‐old male and female genetically obese mice (Ob; n=20) and wild‐type littermates (WT; n=20) were fed a standard diet and randomized to either ad libitum feeding or a 24‐hour fast (WT‐CON, WT‐Fast, Ob‐CON, Ob‐Fast). Mouse weight was measured before and after feeding protocols. Following feeding protocols, arterial stiffness was assessed via arterial pulse wave velocity (aPWV); liver and mesenteric adipose tissue (MAT) immune cell populations were assessed via spectral flow cytometry and normalized by tissue weight. For statistical analyses, sex‐adjusted regression models were fit to account for the confounding effects of sex.

RESULTS

There was not a significant interaction between genotype and intervention on changes in body weight, plasma glucose concentrations, or aPWV (p=0.16; p=0.33; p=0.37, respectively). After adjusting for genotype, acute fasting did not significantly alter changes in plasma glucose concentrations or aPWV (p=0.95; p=0.22). However, after adjusting for genotype, acute fasting promoted significant weight loss (CON 0.4g ± 0.56; Fast ‐2.6g ± 0.72; p<0.01). Compared to Ob‐CON, Ob‐Fast mice had significant increases in liver total immune and cytotoxic T cells (p<0.01) yet no significant changes in either liver regulatory T or helper T cells nor in MAT total immune, cytotoxic T, helper T, and regulatory T cell populations. WT‐Fast mice had significant reductions in liver total immune, cytotoxic T, helper T, and regulatory T cell populations while these same cell populations were increased in MAT when compared to WT‐CON mice.

CONCLUSIONS

An acute 24 hr. fast in mice was insufficient to significantly modulate glucose intolerance and aPWV in Ob mice compared to WT. However, fasting differentially affected liver and MAT T cell populations in mice. These results suggest acute fasting could be a robust mediator of T cell populations with a particular emphasis on expanding liver cytotoxic T cell communities in obese mice.
Automated Evaluation of Respiratory Signals to Provide Insight Into Respiratory Drive
Authors: Khurram Obaid U. , Gransee Heather M. , Sieck Gary C. , Mantilla Carlos B.
Published: May 2022

The diaphragm muscle is the primary inspiratory muscle in mammals and is highly active throughout life. The repetitive activation of the diaphragm muscle (and of other muscles driven by central pattern generator activity) presents an opportunity to analyze these physiological data on a per‐event basis. Here we highlight the development and implementation of the Automated Evaluation of Respiratory Signals (AERS) algorithm to detect individual respiratory events across a range of motor behaviors. Because different respiratory‐related signals provide different types of information, the AERS algorithm is designed to be robust regardless of the signal type—e.g., diaphragm electromyography (EMG) and transdiaphragmatic pressure (Pdi). Individual respiratory event data obtained from adult rats were assessed using the AERS algorithm. This analysis revealed diaphragm root‐mean‐square (RMS) EMG and Pdi were lowest during eupnea, increased slightly in response to hypoxia‐hypercapnia (10% O2 ‐ 5% CO2), and increased approximately 3‐fold in response to airway occlusion compared to the eupneic baseline. The coefficient of variation (CV) within subjects was less than 10% across all behaviors for both RMS EMG and Pdi. By contrast, the CV was approximately 30% across subjects. Respiratory rate was increased by ~20 breaths/min during hypoxia‐hypercapnia compared to eupnea, as expected based on previous work from our lab and others. The respiratory rate CV within subjects was less than 10% during eupnea and hypoxia‐hypercapnia; during airway occlusion, the CV was ~17%, likely owing to the small number of respiratory events (2‐5) analyzed for this behavior per animal. For eupnea and hypoxia‐hypercapnia, the CV across subjects was ~20%, whereas during airway occlusion it was ~25%. Overall, our findings validate the mean values across various measures of respiratory motor output compared to standard subject means approach. However, per‐event analyses greatly enhance statistical power, allowing detection of physiologically relevant measures by taking within‐subject variability into account. Additionally, the AERS analyses show that various measures of respiratory variability as well as time‐frequency analyses can provide deeper insights into respiratory drive when accounting for individual respiratory events. These analyses can be effectively applied to a variety of models of respiratory motor unit loss or dysfunction (e.g., spinal cord injury and aging), allowing the extraction of large amounts of data from respiratory signals for more comprehensive assessments. Our findings suggest that the AERS approach may be particularly beneficial in reducing animal numbers in certain types of studies, for the detection of physiological changes reflected in respiratory signals with interventions causing gain or loss of function, and for analyses of respiratory variability, where breath‐to‐breath dynamics are important.
Integrated Math And Science Scholars (iMASS)‐ Community and collaboration creating opportunities for STEM students
Authors: Kasper Sherry , Schlosser Sarah
Published: May 2022

The Integrated Math and Science Scholars (iMASS) program was developed to support student retention and success in STEM disciplines over the last 6 years. Lee University faculty from 4 different disciplines and a variety of university personnel worked to support 79 students throughout the life of the grant with an emphasis on at risk students including first generation, females, and minority students. It was hypothesized that creating an integrated curriculum for these students and a sense of community through a seminar course we would be able to support and retain more of these students within STEM disciplines. Through innovative curriculum, a weekly seminar course, intentional mentoring, co‐curricular trips, community building activities, and national conference attendance and participation students were given tools and resources to develop professionally. Of the 79 participating students, 35 received scholarships and 44 alternates and associates participated without funding. 61 of the students were retained in STEM at Lee. 32 (91.4%) of the students receiving scholarships were retained in STEM at Lee. The program success also is reflected in some of the larger impacts on the culture of these departments.
Astrocyte‐to‐neuron reprogramming for spinal cord repair
Authors: Niceforo Alessia , Fortino Tara , Shah Yashvi , Zholudeva Lyandysha V. , Qiang Liang , Lane Michael A.
Published: May 2022

Traumatic spinal cord injury (SCI) disrupts neural pathways and results in the devastating tissue damage and loss of function, with subsequent reactive gliosis and tissue scarring that limits further loss, but also limits endogenous repair. One therapeutic strategy gaining some attention in recent years is cellular reprogramming, which harnesses advances in cellular engineering to directly convert cells from one phenotype to another. The present work explores strategies for converting either mature or developing astrocytes into neurons. Our initial experiments have begun exploring viral targeting of glial cells to promote doxycycline‐inducible overexpression of neurogenic transcription factors (TFs) Ascl1 or a combination of microRNAs (miRs) miR124, miR9/9*, for neuronal conversion. Our primary goal was to establish optimal conditions for consistent and efficient reprogramming of developing spinal glia and mature reactive spinal astrocytes.

Primary astrocyte cultures were generated from postnatal days 2‐3 (P2‐3) rat cortex and spinal cord, and then were activated with TGFβ (Transforming growth factor beta) one day prior to viral transduction. Fifteen days following doxycycline administration, astrocytes took on a typical neuronal morphology. Immunocytochemistry confirmed that these cells were positive for neuronal markers MAP2, NeuN and TAUR1, and negative for GFAP. Multielectrode array recordings of converted cells revealed spontaneous firing consistent with neuronal activity.

These preliminary results confirm that cortical and spinal astrocytes can be efficiently reprogrammed into neurons by either Ascl1 or miRs. Advances in cellular reprogramming are providing novel ways to manipulate the cytoarchitecture of the injured cervical spinal cord. Our ongoing work is now using this same approach to modify developing glial precursors that can be harness for their pro‐reparative properties for spinal cord repair.
Investigating the tryptophan‐metabolizing enzyme AFMID (arylformamidase) in colon cancer
Authors: Garcia Roy , Conacci Sorrell Maralice
Published: May 2022

Tryptophan is an essential amino acid which is metabolized into catabolites necessary for physiological homeostasis. Of the various catabolic fates of tryptophan, the kynurenine pathway is of both basic and clinical interests due to its signature catabolite kynurenine which contributes to immune privilege, is a precursor of NAD +, and acts as an activating ligand of the transcription factor aryl hydrocarbon receptor (AHR.) Through elevated expression of tryptophan transporters and tryptophan‐catabolizing enzymes, cancer cells promote the production of kynurenine and downstream catabolites. Oncogene‐mediated upregulation of tryptophan transporters and tryptophan catabolizing enzymes IDO1, TDO2, and arylformamidase (AFMID) occur in several cancer types including breast, esophageal, liver, and colon cancers with correlation to poor prognosis. Targeting of the kynurenine pathway has therefore been a clinical focus; however, all initial clinical trials aiming to inhibit IDO1 failed. A new generation of clinical trials targeting IDO1, TDO2, and tryptophan transporters are underway, yet no preclinical evaluation of the enzyme (AFMID) has ever been conducted. AFMID acts immediately downstream of IDO1, IDO2, and TDO2 to catalyze the reaction that produces kynurenine itself. Unlike IDO1/2 and TDO2 which are functionally redundant, AFMID features a unique function. Despite its importance, no studies have comprehensively evaluated AFMID’s direct roles in metabolic reprogramming, cell‐autonomous proliferation, or microenvironment remodeling. We aim to investigate the cell autonomous and immune modulatory functions of AFMID in colon cancer and to define novel approaches to inhibit AFMID activity as strategy to block the kynurenine pathway.
Regional and Single Nucleotide Correction of Sequence Bias in Chromatin Accessibility Data
Authors: Wolpe Jacob , Guertin Michael
Published: May 2022

Chromatin accessibility assays directly measure which regions of DNA are actively regulated in the genome. Genomic chromatin accessibility data provide the gold standard of spatial and temporal resolution of regulatory landscapes in various cell types and conditions. Accessibility assays depend on enzymatic digestion of DNA and these enzymes are biased with respect to the sequences they recognize and cut. Sequence biases can lead to incorrect conclusions about the nature of the chromatin landscape within an experiment. Previous work from our group has shown that we can use observed k‐mer cut frequencies and k‐mer representation in the genome to correct the majority of this bias. However, these k‐mer scaling approaches did not effectively correct ATAC‐seq data, which assesses chromatin accessibility using the Tn5 transposase enzyme. Here, we find that the oligomeric state of Tn5 and wide sequence bias window precludes effective k‐mer scaling to account for Tn5 sequence bias. Tn5 sequence specificity is influenced by sequence content up to ten bases away from the Tn5 insertion site in both directions. In comparison, Benzonase, Cyanase, MNase, and DNase are primarily influenced by the sequence within 3 bases of the cleavage site. We could not effectively apply k‐mer scaling to ATAC‐seq data because the majority of insertion sites happen at unique 20‐mers in the genome and the method requires many genomic instances per k‐mer. To overcome this limitation, we applied an interpretable machine learning approach, prediction rules ensemble, to correct Tn5‐digested naked DNA. Application of a prediction rules ensemble effectively corrects local (single nucleotide resolution) sequence bias as well as sequence biases influenced by regional GC content. Correction of Tn5 sequence biases is important as the field continues to generate ATAC‐seq data at accelerating rates and use these data to accurately infer regulatory element activity.
The 2021 Marine Pharmacology and Pharmaceuticals Pipeline
Authors: Wu Alex C. , Jelielek Karolina K. , Le Henry Q. , Butt Maryam , Newman David J. , Glaser Keith B. , Pierce Marsha L. , Mayer Alejandro M.
Published: May 2022

Objective

Marine organisms continue providing significant chemical biodiversity that contributes to both the global preclinical and clinical pharmaceutical pipelines. A recently (2021) redesigned website www.marinepharmacology.org allows for easier and more frequent updates, thus highlighting the continuous progress in this area of marine pharmacology/pharmaceutical research and drug development.

Hypothesis

There has been progress in the approval and development of marine‐derived drugs in 2021.

Methods

The following four websites were researched to determine new marine‐derived drugs approved in 2021: (a) USA: https://www.accessdata.fda.gov/scripts/cder/daf/ United States Food and Drug Administration (FDA); (b) Europe: https://www.clinicaltrialsregister.eu/ctr‐search/search European Clinical Trials register; (c) China: https://www.chictr.org.cn/enindex.aspx Chinese Clinical Trials register; and (d) www.clinicaltrials.gov to determine the current status of marine‐derived compounds in Phase I, Phase II, and Phase III of drug development.

Results

Our research supports our hypothesis because in 2021 there has been continued progress in the approval and development of marine‐derived drugs. In 2021 the clinical marine pharmaceutical pipeline consisted of 15 marine‐derived drugs approved by the FDA, 2 approved in Australia, and 1 approved in China: for cancer, cytarabine (Cytosar‐U®, Depocyt®, FDA‐approved 1969); for Herpes Simplex Virus, vidarabine (Vira‐A®, FDA‐approved 1976); for pain, ziconotide (Prialt®, FDA‐approved 2004); for hypertriglyceridemia, omega‐3‐acid ethyl esters (Lovaza®, FDA‐approved 2004, Vascepa®, FDA‐approved 2012, Epanova®, FDA‐approved 2014); for cancer, eribulin mesylate (Halaven®, FDA‐approved 2010), brentuximab vedotin (Adcetris®, FDA‐approved 2011), trabectedin (Yondelis®, FDA‐approved 2015), panobinostat (Farydak®, FDA‐approved 2015), plitidepsin (Aplidine®, approved in Australia 2018), polatuzumab vedotin (Polivy™, FDA‐approved 2019), enfortumab vedotin‐ejfv (PACDEV™, FDA‐approved 2019), lurbinectedin (Zepzelca™, FDA‐approved 2020, Australia 2021), belantamab mafadotin‐blmf (Blenrep™, FDA‐approved 2020), disitamab vedotin (Aidixi™, approved in China 2021), and tisotumab vidotin (Tivdak™, FDA‐Approved 2021). Furthermore, we determined that there were 33 marine‐derived compounds in active clinical trials in 2021: 5 marine‐derived compounds were in Phase III, 12 compounds in Phase II, and at least 16 compounds in Phase I. Many of compounds classified as auristatin‐containing antibody drug conjugates.

Conclusions

In 2021, new marine‐derived drugs were approved by FDA, as well as in Australia and China. The clinical development of marine‐derived drugs remained very robust and active with several advancing to Ph III, Ph II and entering at Ph I.
A combinatorial chemical epigenetics screen identifies an off‐target modulation of drug transporter function
Authors: Barghout Samir H. , Yu Yifan , Arrowsmith Cheryl H. , Barsyte‐Lovejoy Dalia
Published: May 2022

Anticancer drug response is determined by genetic and epigenetic mechanisms. To identify the epigenetic regulators of anticancer drug response, we conducted a chemical epigenetics screen using chemical probes that target different epigenetic modulators. In this screen, we tested 31 epigenetic probes in combination with 14 mechanistically diverse anticancer agents and measured the viability in A549 lung adenocarcinoma cells by the Resazurin assay. We identified 6 epigenetic probes that significantly potentiated the cytotoxicity of TAK‐243, a first‐in‐class ubiquitin‐activating enzyme (UBA1) inhibitor evaluated in several solid and hematologic malignancies. These probes include TP‐472, GSK‐864, A‐196, UNC1999, SGC‐CBP30 and PFI‐4, and target BRD9/7, mutant IDH1, SUV420H1/H2, EZH2/H1, p300/CBP and BRPF1B, respectively. To validate the screen results, we assessed the viability after combination of TAK‐243 and the identified probes and observed 4‐ to 30‐fold potentiation of TAK‐243 cytotoxicity in myeloma cells. Moreover, we tested the identified probes in 13 additional cell lines and observed profound potentiation. Upon combination with a panel of anticancer agents, epigenetic probes did not potentiate their cytotoxicity suggesting the observed effects are selective for TAK‐243. In contrast to epigenetic probes, negative chemical controls did not have a significant impact on TAK‐243 cytotoxicity. As assessed by immunoblotting, potentiation of TAK‐243 cytotoxicity was associated with reduced ubiquitylation and induction of apoptosis. Using the cellular thermal shift assay (CETSA), UBA1 displayed increased engagement with TAK‐243, suggesting the epigenetic probes enhanced intracellular drug accumulation. Mechanistically, the epigenetic probes exerted their potentiation by inhibiting the efflux transporter ABCG2 without inducing significant changes in ubiquitylation pathways or ABCG2 expression levels. Additionally, the identified probes shared significant chemical scaffold similarities with TAK‐243. Based on the screen results, we developed a cell‐based assay that exploits TAK‐243 and ABCG2‐overexpressing cells to reliably quantify the ABCG2‐inhibitory activity of novel compounds. In conclusion, our study identifies epigenetic probes that profoundly potentiate TAK‐243 cytotoxicity through off‐target ABCG2 inhibition. We also provide experimental evidence of the inability of negative controls to exclude a subset of off‐target effects of chemical probes. Finally, we have developed a robust cell‐based assay that can quantitatively evaluate ABCG2 inhibition by drug candidates.
Transcriptomic Analysis of Peripheral Artery Disease Patient Derived Myotubes Reveals Broad Gene Expression Changes and Alternative Splicing
Authors: Ring Andrew , Ismaeel Ahmed , Fletcher Emma , Papoutsi Evlampia , Miserlis Dimitrios , Koutakis Panagiotis
Published: May 2022

Peripheral Artery Disease (PAD) is a disease characterized by atherosclerosis of the arteries suppling blood to the lower extremities. One of the clinical manifestations of PAD is myopathy of the affected muscles. In order to elucidate the role of gene expression changes in myotubes and their effect on PAD myopathy, we isolated myotubes from PAD and control patients to perform transcriptomic analysis. We hypothesized that myogenic and muscle contractility related gene expression levels change during PAD. Our analysis revealed 690 significant differentially expressed genes. Gene ontology analysis showed significant increase in genes relating to myogenesis and muscle contraction. Hypoxia and mitochondria related genes were also dysregulated in PAD myotubes. Kegg pathway analysis showed genes enriched in muscle contraction pathways as well as adrenergic signaling. Alternative splicing analysis showed 209 genes with differential transcript isoform expression. Gene ontology showed that these genes relate to cell adhesion, apoptosis and autophagy. qPCR and Protein expression analysis confirms some of these transcriptional changes. This study shows the need for further exploration into how transcriptional changes affect the development of myopathies and the progression of PAD. It also shows the ability for isolation patient myotubes as a model for the study of myopathies in PAD.
The Single Dose Cocaine Reinstatement of Lever‐Pressing Behavior in Rats is Explained by The Compulsion Zone Theory
Authors: Desai Jhanvi N. , Zinani Dakota B. , Meyer Nicole J. , Norman Andrew B.
Published: May 2022

The compulsion zone theory of the cocaine self‐administration paradigm states that lever‐pressing behavior is induced when cocaine levels fall to or below the satiety threshold. Single injections of cocaine are known to reinstate lever‐pressing in rats trained to self‐administer cocaine. We investigated whether this phenomenon can be explained by the compulsion zone theory. Male Sprague‐Dawley rats (n = 5) were trained to self‐administer cocaine i.v. on an FR‐1 schedule. Once trained, these rats were administered a single dose of cocaine (2 – 12 µmol/kg, i.v.) in four daily sessions per week, and the time to lever‐pressing, the number of presses and the duration of pressing were recorded. The latency from the start of an injection to the resumption of lever pressing increased with the cocaine dose and ranged from approximately 2 min at 2 µmol/kg to more than 15 min at 12 µmol/kg. The D1 dopamine receptor antagonist SCH23390 (10‐30 nmol/kg i.v.) decreased the latency to reinstatement of lever pressing, following a dose of 12 µmol/kg cocaine. The increasing latency to lever‐pressing is consistent with the increasingly higher cocaine levels taking longer to fall to the satiety threshold, while the lower doses are immediately within the compulsion zone. An antagonist‐induced elevation of the satiety threshold explains the reduced latency to reinstatement. The number and duration of lever presses may reflect the time that the cocaine levels are within the compulsion zone. The cocaine satiety threshold model explains the dose‐dependent delay in reinstatement of lever‐pressing behavior and the effect of dopamine antagonists.
Utilization of glucan phosphatase Starch Excess4 to enhance in‐vitro starch degradation
Authors: Weis Kenyon , Raththagala Madushi
Published: May 2022

Starch is one of the most attractive natural polymers because of its biodegradability, abundance, and renewability. However, the inert nature and semicrystalline structure of native starch granules make starch essentially unusable for most industrial applications without further processing. Therefore, starch is additionally modified via physical, chemical, and biotechnological methods to increase processability and function. Starch obtained from corn, rice, wheat, potato, and cassava, serves as the main resource for the majority of starch‐based products. However, much of our understanding of starch metabolism is coming from Arabidopsis thaliana transitory starch metabolism. More systematic studies are needed to understand starch turnover in different species and organs of biotechnologically important species. Reversible starch phosphorylation is essential for efficient starch degradation and SEX4 is a key enzyme in this process. We employed biochemical and functional analyses to study SEX4 from agronomically important crops. Taken together, our findings provide an in depth understanding of SEX4 dephosphorylation of agronomically important plants and green algae species and provide compelling evidence of how SEX4 can be utilized to enhance native starch degradation.
Human transcription factor protein interaction networks
Authors: Varjosalo Markku , Göös Helka , Kinnunen Matias , Salokas Kari , Liu Xiaonan
Published: May 2022

Transcription factors (TFs) interact with several other proteins in the process of transcriptional regulation. Here, we identify 6703 and 1536 protein–protein interactions for 109 different human TFs through proximity‐dependent biotinylation (BioID) and affinity purification mass spectrometry (AP‐MS), respectively. The BioID analysis identifiesmore high‐confidence interactions, highlighting the transient and dynamic nature of many of the TF interactions.

By performing clustering and correlation analyses, we identify subgroups of TFs associated with specific biological functions, such as RNA splicing or chromatin remodeling. We also observe 202 TF‐TF interactions, of which 118 are interactions with nuclear factor 1 (NFI) family members, indicating uncharacterized cross‐talk between NFI signaling and other TF signaling pathways. Moreover, TF interactions with basal transcription machinery are mainly observed through TFIID and SAGA complexes.

This study provides a rich resource of human TF interactions and also act as a starting point for future studies aimed at understanding TF‐mediated transcription.
Effects of High Frequency Resistance Training on RNA Expression Related to Inflammation: A Case Report
Authors: Saffold Keith , Kelley Christian , Buford Thomas
Published: May 2022

BACKGROUND

Resistance training (RT) is the main modality of exercise used to induce hypertrophic and neuromuscular adaptations. RT done at high intensities (HI‐RT) is also capable of inducing epigenetic modifications leading to altered protein expression. HI‐RT increases protein expression of many enzymes that play key roles in disease prevention and treatment. Inflammation is one pathology that is associated with many disease states that can be altered by the effects of HI‐RT. Both pro‐ and anti‐inflammatory cytokines are upregulated after a bout of exercise. Long term training leads to increases in anti‐inflammatory cytokines such as muscle‐derived interleukin‐6 (IL‐6). Current recommendations state that each muscle group should be exercised 2‐3 nonconsecutive days per week to allow time for recovery. HI‐RT protocols including training much higher training volumes and frequencies have been used by advanced athletes for decades, the effects of these protocols on health outcomes are not reflected in the literature.

STUDY OBJECTIVE

The objective of this case report is to determine the effects of high‐frequency and high‐volume HI‐RT on RNA expression linked to inflammatory cytokines.

HYPOTHESIS

It is hypothesized that both the pro‐inflammatory and anti‐inflammatory cytokines will be upregulated following a high‐frequency high‐volume HI‐RT program due to increased muscle damage and macrophage infiltration.

METHODS

The participant (N = 1; 24 years; 192 cm; 107.7 kg; 25.6% body fat) had leg circumference measured at 50% of the length of the femur. Then, a muscle biopsy was performed on the right leg vastus lateralis (VL). Muscle samples were stored at ‐80°C for later analysis. The participant completed a high‐frequency, high‐volume HI‐RT program for one month, which included barbell squats 5 days per week. After the training program was completed, leg circumference was measured again and a second biopsy of the right leg VL was taken. RNA was isolated from the samples for real‐time polymerase chain reaction (RT‐PCR) analysis of IL‐6 and tumor necrosis factor alpha (TNF‐α).

RESULTS

The participant experienced a 4 cm increase in thigh circumference (6.4%). Due to time constraints, RT‐PCR analysis was delayed. Analysis will be concluded prior to the conference meeting.

CONCLUSIONS

To be updated after analysis is complete.
The Effects of Morning vs. Evening Mindfulness Meditation on Sleep, Anxiety, and Decentering: A Pilot Analysis
Authors: Basala Thomas , Morin Brigitte , Durocher John
Published: May 2022

According to the CDC, most children and about 1 in 3 adults in the United States fail to reach their age group recommended quantity of sleep. Recently, non‐pharmacological sleep aids, such as meditation, have gained popularity. However, to our knowledge there is no research data that compares sleep metrics or psychological well‐being between those that meditate in the morning vs. in the evening. This study aims to investigate the potential circadian‐dependent effects of meditation timing and how it may influence metrics of sleep and psychological well‐being. We hypothesized that those who meditate just before sleep would have improved sleep parameters and self‐reported feelings of psychological well‐being when compared to those that meditate in the morning. Sixteen college students participated in the study, eight who meditated in the morning (5M; 3F) after waking and eight who meditated in the evening (4M; 4F) just before going to sleep. Participants were asked to wear an Actiwatch Spectrum PRO to evaluate physical activity and sleep throughout the recording periods. The first phase of the protocol consisted of a 4‐day (Monday‐Thursday) baseline recording period where total sleep time (TST) and sleep efficiency measures were recorded. Participants were instructed to not meditate during the baseline period. During the 4‐day intervention period which occurred the following week (Monday‐Thursday), participants were randomized to a 24‐minute pre‐sleep or morning mindfulness meditation intervention from the app Insight Timer. At the conclusion of both baseline and intervention time periods (on Friday of each respective week), participants were instructed to complete a state‐trait anxiety inventory (STAI), a Five Facts of Mindfulness Questionnaire, and a Decentering Questionnaire. Means were considered to be significantly different when p<0.05. The acute meditation protocols did not significantly change TST from baseline in the morning (6.7±0.2 vs. 6.8±0.2 hours) or evening (6.9±0.3 vs. 7.2±0.2 hours) meditators. Likewise, sleep efficiency was not significantly changed in either group. However, there was a significant decrease in both state and trait anxiety (baseline vs. treatment, p<0.02 for both), and a significant increase in the five facets of mindfulness and ability to decenter (baseline vs. treatment, p<0.01 for both). Our preliminary results suggest that acute meditation can help to improve several measures of psychological well‐being whether it is done in the morning or evening. Further investigation within our own study and from others may help to better understand whether evening meditation can offer specific sleep benefits.
Metabolic and Transcriptomic Effects of Mediator Kinase Inhibition on the Interferon Response in Down Syndrome
Authors: Cozzolino Kira A. , Jones Taylor , Ajit Deepa , Dowell Robin , Taatjes Dylan
Published: May 2022

Down syndrome, also known as Trisomy 21, is a condition in humans resulting from full or partial triplication of chromosome 21. Affected individuals present with a constellation of symptoms that includes an increased risk of autoimmune disorders. Cells of patients with Down syndrome exhibit chronic, hyperactivated interferon (IFN) signaling, due in part to the presence of several interferon receptors on chromosome 21. Partial knockout of these receptors has been shown to increase cellular viability, indicating that treatments targeting the IFN response may ameliorate the comorbidities associated with Down syndrome.

Among the regulators of the IFN response are the Mediator‐associated kinases, CDK8 and CDK19. Studies using inhibitors such as cortistatin A, which targets the enzymatic activity of both kinases, have demonstrated improved outcomes in autoimmune disease models. However, the specific mechanisms and contributions of CDK8 vs. CDK19 remain unclear. In mouse and human cells, we and others have shown that the transcriptional response to IFN is different when CDK8/19 are inhibited as opposed to when CDK19 specifically is degraded, suggesting that each kinase regulates distinct sets of IFN‐responsive genes through divergent mechanisms. While it seems likely that CDK8 acts through phosphorylation of proteins such as the transcription factor STAT1, CDK19 may instead serve as a physical scaffold.

To begin to examine whether inhibition of the Mediator kinases is a viable strategy to treat Down syndrome‐associated comorbidities, I assessed the metabolic effects of CDK8/19 inhibition in patient‐derived lymphoblastoid cell lines from a pair of age‐ and sex‐matched siblings, one with Down syndrome (T21) and one without (D21). Metabolomic analysis demonstrated a severely blunted response to IFNγ treatment in T21 cells as compared to D21 cells, and revealed genotype‐specific changes in energy, nucleotide, and fatty acid metabolism. Interestingly, a subset of metabolites that were decreased or elevated in T21 cells treated with IFNγ alone showed no significant changes in cells also treated with cortistatin A, suggesting that Mediator kinase inhibition may reduce the responsiveness of T21 cells to IFNγ.

Ongoing RNA‐seq experiments are assessing the transcriptional response to IFN in D21 and T21 cells following CDK8/19 inhibition, with a focus on potential splicing defects and altered gene expression programs in T21. In sum, these experiments will address the under‐explored question of how T21 alters metabolic and transcriptional responses to IFNγ and how these responses are controlled by the Mediator kinases. This work will also shed light on the therapeutic potential of Mediator kinase inhibitors, as a means to dampen hyper‐activated IFN signaling in Down syndrome.