bluuebunny/crossref_metadata_2025 huggingface dataset:
Mitochondrial receptor NIX meidates mitophagy and protects HeLa cells in against Cdinduced toxicity.
Authors: Naeem Sajid
Published: April 2020

Heavy metals exposure are coupled with cellular damages. Our previous studies have demonstrated induction of ROS dependent mitochondrial damages and decreased cell viability in response to heavy metals exposure. Damaged mitochondria are selectively removed by PINK1/Parkin pathway in response to heavy metal exposures. Moreover, there exist several other mitophagy pathways, which are activated under various conditions and may crosstalk with each other. In present study we used HeLa cells which have mutated non‐functional Parkin, thus unable to conduct Parkin‐mediated mitophagy, to evaluate the role of other mitophagy pathways induced by Cd. Our results showed that Cd exposure could induce mitophagy in HeLa cells, lacking Parkin, and GM00637 cells, having intact Parkin, which was evaluated by translocation of mitochondria to lysosomes and decreased mitochondrial markers. In GM00637 cells, the mitophagy process was mediated by PINK1/Parkin pathway as confirmed by noticing Parkin translocation to mitochondria. However, in HeLa cells, no Parkin was detected. Interestingly, NIX expression was significantly increased in HeLa cells exceptionally. Moreover knocking down of NIX in HeLa cells resulted in decreased cell viability and inhibition of mitophagy process in HeLa cells. Finally, Cd exposure significantly increased NIX phosphorylation at serine‐81. These finding suggest a compensatory role of NIX in heavy metals mediated mitophagy when Parkin is not present.
CKDNET: a program for fighting with chronic kidney disease in the Northeast Thailand
Authors: Cha’on Ubon , Pongskul Cholatip , Thinkhamrop Bandit , Limwattananon Chulaporn , Sharma Amod , Anutrakulchai Sirirat
Published: April 2020

Background

Chronic kidney disease (CKD) in the Northeast Thailand (22.2%) was in the highest range of the world, therefore, Chronic Kidney Disease Prevention in the Northeast of Thailand (CKDNET) program was established to prevent and reduce CKD in this area, https://ckd.kku.ac.th/.

Methods

CKDNET program attempt to cover all general population, high risk groups and all stages of CKD patients with several strategies including early screening, effective CKD registry, CKD comprehensive care models in community and hospital levels, as well as cost effectiveness analysis.

Results

Preliminary results from CKD screening in general population of two rural sub‐districts shows that the prevalence of CKD is approximately 27%. Ten million individuals are registered in CKD hospital registry with hundred thousands of CKD cases. The CKD guideline that suits for local situation has been formulated and CKD care trainings were developed for physicians and nurses. Moreover, our multidisciplinary intervention and self‐management supports were effective to improve glomerular filtration rate and blood pressure, and quality of life among CKD patients.

Conclusion

CKDNET program is the holistic approach to deal with the different aspects of CKD which in turn to attenuate the CKD burden in the Northeast Thailand.

Support or Funding Information

Faculty of Medicine, Khon Kaen University, Thailand
The Role of Direct Effects of Drugs of Abuse Binding to the hDAT Molecule in their Diverse Behavioral Phenotypes
Authors: Bergman Shana , Razavi Asghar , Khelashvili George , Weinstein Harel
Published: April 2020

Substrates, inhibitors, and various classes of drugs of abuse that target the human dopamine transporter (hDAT) elicit a variety of responses measurable in vitro and in vivo. Classification of such ligands in terms of their chemical and/or pharmacological properties has failed to produce sufficient mechanistic explanations, or hypotheses, to guide a satisfactory understanding of the nature of this diversity in the effects of hDAT ligands. With the objective of advancing the current understanding of these drugs and their mechanisms of action, we set out to investigate the role that the response of the hDAT molecule to the binding of these ligands may have in their known diverse behavioral effects. We use structurally and pharmacologically diverse ligands of hDAT (e.g., typical inhibitors, atypical inhibitors, substrates and releasers) in extensive all‐atom molecular dynamics (MD) simulations to reveal the structural and dynamic effects that characterize the response of the transporter molecule to each of them and to enable a classification of hDAT molecular responses. MD trajectories describing the time‐dependent dynamic behavior of each complex are analyzed to quantify the molecular responses of hDAT, and machine learning algorithms serve to classify each response and reveal specific dynamic features underlying the molecular mechanism of hDAT’s response to each ligand. This approach offers mechanistic insight into the role of the ligand‐determined molecular response of hDAT in determining the diversity of effects expressed in the functional outcomes.

Support or Funding Information

SB is supported by T32 “Genetic and environmental influences on addiction” (DA03980), research is supported by NIH Grant R01 DA041510 (to H.W.)
Novel FRET‐based biosensors for monitoring Presenilin/γ‐secretase activity in live cells
Authors: Maesako Masato , Sekula Nicole M. , Anderson Lauren C. , Berezovska Oksana
Published: April 2020

Presenilin (PS)/γ‐secretase is responsible for the proteolytic processing of wide variety of membrane associated proteins that include APP and Notch. Therefore, a change in PS/γ‐secretase activity is linked to essential biological events as well as to the progression of many diseases. However, not much is known about how PS/γ‐secretase activity is spatiotemporally regulated in live cells, mainly because shortcomings of the existing assays prevent monitoring dynamic changes in the activity of PS/γ‐secretase in its normal physiological environment. Here we present successful development and validation of the Förster resonance energy transfer (FRET)‐based biosensors, so called the APP C99 Y‐T and the Notch1 N100 Y‐T biosensors, that enable quantitative monitoring of endogenous PS/γ‐secretase activity in live cells longitudinally on a cell‐by‐cell basis. Using these novel FRET biosensors, we uncovered dynamic spatiotemporal pattern of PS/γ‐secretase activity that is heterogeneously regulated among live neurons.

Support or Funding Information

NIH: AG 44486 (OB), AG 15379 (OB) and BrightFocus Foundation: A2019056F (MM)
Amino Acid Complexed Zinc Has Higher Bioavailability Than Hydroxy Acid Complexed Zinc
Authors: Crabtree Amber , Shay Sheila , Moore Christie , Gladson Santhi , West James
Published: April 2020

Poor zinc absorption causes zinc deficiency, leading to skin, gut, and immune pathologies, and psychiatric changes, and are potentially life threatening in infants and children. This can be caused by dietary deficiencies, but also inherited mutations of the ZIP4 gene.

Objective

Although zinc complexed with organic molecules have previously been shown to have better uptake, we hypothesized that bonding the zinc to the amino group would be less liable to breakdown in the gut than zinc complexed to hydroxy acids, leading to higher bioavailability.

Methods

We treated wild‐type and mice with inducible universal knockout of one copy of the ZIP4 gene with reduced zinc diet (15 ppm) ad libitum, either in the form of amino acid complexed (CSK18271) or hydroxy acid complex (CSK18273) zinc in an otherwise nutritionally complete diet. Weight was measured at weekly intervals, and mice placed in metabolic phenotyping chambers for a week near the end of the trial.

Results

We found that ZIP4+/− mice fed CSK18271 had weight gain comparable to wild‐type mice; wild‐type mice fed CSK18273 had lower weight gain, indicating bioavailability was lower even with intact zinc transporter. Zip4+/− mice fed CSK18273 had the worst overall weight gain. Metabolic phenotyping chambers found an increase in respiratory exchange rate (RER) and water intake in the Zip4+/− mice, indicating reduced kidney function in the heterozygous knockout mice, which was compensated for functionally by CSK18271 diet.

Conclusions

Zinc complexed to the amine group has higher bioavailability than zinc complexed to the hydroxyl group in amino acids, likely because of increased resistance to degradation in the gut. This results in improved weight gain in both wild‐type and Zip4+/− animals.

Support or Funding Information

The work was funded by Vanderbilt University Medical Center internal funds.
TGFBI inhibits endothelial angiogenesis stimulated by adipocyte conditioned medium.
Authors: Lee Seul Gi , Nam Ju-Ock
Published: April 2020

Several studies have suggested that the ECM remodelling or environment is tightly related to the adipogenesis and adipose angiogenesis. In the present study, our data showed that TGFBI (transforming Growth Factor‐Beta–Induced) suppressed angiogenesis induced by adipocyte conditioned medium (CM) in vitro and in vivo. Treatment of TGFBI protein (TGFBIp) diminished the migration and tube‐forming capacity of SVEC‐10 endothelial cells cultured in CM. In contrast, TGFBI knockout (KO) mice contained an increased number of vessels and infiltration into matrigel containing CM. By mechanism of action, TGFBI regulated cleavage and activation of focal adhesion protein (FAK) in adipose tissue. The altered FAK cleavage by TGFBI may account for the modulation of endothelial cell function ultimately impacting adipose angiogenesis. Altogether, these findings demonstrate the potential role of TGFBI in controlling endothelial angiogenesis induced by adipocytes.
Effects of white adipose tissue denervation on cardiovascular and renal parameters in Wistar rats treated with high‐fat diet
Authors: Nishi Erika E. , Ferreira Giovanna R. , Garcia Michele L. , Campos Ruy R. , Bergamaschi Cassia M. T.
Published: April 2020

Sympathetic nervous system and renin‐angiotensin‐aldosterone system overactivation plays a major role in cardiovascular changes in obesity. The white adipose tissue (WAT) is an essential endocrine organ that synthesizes several biologically active compounds that regulate metabolic homeostasis and contains both sensorial and sympathetic innervation. This study aimed to investigate the effects of retroperitoneal white adipose tissue (rWAT) denervation (DNX) on cardiovascular and renal parameters in obese rats. Animals were treated with a high‐fat diet (HFD) for 8 weeks to induce obesity and underwent surgical DNX procedure at 6 weeks. At the end of the experimental protocol, animals treated with HFD were significantly overweight, whereas the HFD+DNX group had a slightly decrease in body weight. The HFD group presented a discrete increase in baseline mean arterial pressure, which was normalized after DNX. Moreover, DNX improved bradycardic response gain when cardiac baroreflex control was evaluated in the HFD group. No changes were found in renal parameters. However, it was found a significant decrease in circulating angiotensinogen and leptin after rWAT DNX. Therefore, these findings suggest that rWAT innervation has influence on cardiovascular changes found in obesity.

Support or Funding Information

CAPES, CNPq and FAPESP
Flip it or not for histology teaching in dental school?
Authors: Sun Xinxin , Leavis Paul
Published: April 2020

Over the years, many novel electronic teaching tools and approaches have complemented and supplanted traditional educational offerings in histology such as lectures. The teaching method frequently used in our dental school histology class is the “flipped classroom” model. Prior to formal classroom sessions, students preview course materials from videos and the syllabus. During formal classroom sessions, students apply the knowledge that they acquired before classes by answering quiz questions using the interactive electronic tool, Poll Everywhere. The instructors will see the live responses from students and review important concepts in histology. The flipped classroom has changed the traditional teacher‐centered lecture‐based teaching method and is designed to encourage self‐motivated learning. However, the attendance of the flipped classroom is very low, even though the interactive class is not recorded.

To find a better strategy for teaching histology in dental school, we compared the effectiveness of “flipped classroom” with a non‐flipped interactive learning class. In the non‐flipped classroom, dental students are not required to watch videos before class. Instead, the instructor provides a brief introduction of day’s topic in class, then students will practice their understanding of freshly learned knowledge by discussing and answering quiz questions using the same interactive tool, Poll Everywhere. Since, in this model, the interactive classes are recorded, students may choose to forgo attending the scheduled interactive classes.

In this study, we used the flipped class model for D22 students and the non‐flipped interactive approach for D23 students. The lecture content, quiz questions, instructor and exam questions were identical for the two classes. We compared the normalized exam performances of students who attended class in the two models.

The research results showed that dental students who were exposed to the non‐flipped interactive class have significantly higher attendance and better normalized exam performance than those who received the flipped class. In the flipped classroom model, dental students exhibit a strong tendency to use the pre‐class videos and syllabus only, which is basically a traditional lecture‐based teaching method without scheduled time restrictions. In contrast, the non‐flipped interactive class model encourages students to attend the classes, even if they missed reviewing pre‐class materials (for example, syllabus). Once the students come to the classroom, they will benefit from the interactive learning that follows the instructor’s outline of the topic.

This study will provide insights for choosing histology teaching strategies for dental students.

Support or Funding Information

Department of Medical Education, Tufts University, School of Medicine; Dental Research Administration, Tufts University School of Dental Medicine
Endurance Exercise Training Does Not Alter Key Receptors Within the Renin‐Angiotensin System in Skeletal Muscle
Authors: Nguyen Branden , Deminice Rafael , Yoshihara Toshinori , Lawrence Jensen , Hyatt Hayden , Ozdemir Mustafa , Powers Scott
Published: April 2020

Prolonged skeletal muscle inactivity results in muscle fiber atrophy and weakness. Interestingly, endurance exercise training promotes a protective phenotype in skeletal muscle fibers that resists inactivity‐induced muscle wasting. The mechanism(s) responsible for this exercise‐induced preconditioning of muscle remain unknown and are explored in these experiments. In this regard, evidence reveals that activation of the classical arm of the renin‐angiotensin system (RAS) contributes to inactivity‐induced muscle wasting. Specifically, activation of the classical RAS pathway via the angiotensin II type I receptor (AT1R) is associated with activation of proteases and muscle fiber atrophy. Additionally, a non‐classical RAS axis exists that acts antagonistically to the classical RAS axis. Therefore, activation of the non‐classical RAS pathway via the Mas receptor (MasR) inhibits signaling through the classical RAS pathway. Indeed, pharmacological studies reveal that blockade of the AT1R and/or activation of the non‐classical RAS axis protects skeletal muscle against inactivity‐induced muscle atrophy. Studies also reveal that endurance exercise training decreases the abundance of AT1R and increases the quantity of MasR in blood vessels. If similar changes occur in skeletal muscles, these changes in receptor abundance could shift the RAS signaling to favor the non‐classical RAS axis and reduce activation of classical RAS signaling in muscle fibers during prolonged inactivity. These experiments test the hypothesis that endurance exercise training decreases the abundance of AT1R and increases the abundance of MasR in skeletal muscle fibers. To test this postulate, adult rats performed 10 days of endurance exercise training (60 min/day @ ~70% VO2Max). At the completion of the training protocol, the abundance of both AT1R and MasR were determined in isolated single muscle fibers. Moreover, a sensitive angiotensin II (Ang‐II) binding assay was employed to confirm exercise‐induced changes in AT1R abundance in muscle. The results reveal that endurance exercise training does not alter the protein abundance of either the AT1R or MasR in the soleus, plantaris, or diaphragm muscle fibers. Moreover, exercise training did not change the Ang II‐binding affinity within skeletal muscles. Collectively, these results do not support the hypothesis that endurance exercise training results in a decrease in the abundance of AT1R and an increase in MasR in skeletal muscle fibers. It follows that exercise‐induced changes in the abundance of RAS receptors does not appear to be the mechanism responsible for exercise‐induced protection of muscle fibers against inactivity‐induced muscle atrophy.

Support or Funding Information

NIH AR064189 (SKP).
GAME BOARD IMPROVES THE ENDOCRINE SYSTEM ANATOMY TEACHING AND LEARNING PROCESS
Authors: de Paula Uchôa Francisco Ewerton , Nunes Bruna Marques Barreto , Forte Guilherme Aguiar , Filho Cézar Nilton Rabelo Lemos , Cerqueira Gilberto Santos , Morano Domingos Antonio Clemente Maria Silvio
Published: April 2020

Board games are important tools of active methodology in the anatomy teaching‐learning process, especially when traditional learning is to be diminished. Several studies demonstrate that board games are effective in promoting learning. At the present time, in our University, the traditional methodology is predominant, and building an appropriate and qualified learning environment is a big challenge nowadays. Based on these assumptions, the aim of this paper was to investigate whether the endocrine anatomy board game promotes meaningful learning. A descriptive exploratory study with quantitative approach was conducted with 31 students of the discipline of anatomy. For learning assessment, a tested and validated board game of endocrine anatomy was applied. For statistical analysis, the normality of the data was verified and the Mann‐Whitney test and the Chi‐Square test were used, and those with p <0.05 were considered significant. It was observed that there is a statistically significant difference between the ages of the participants, and females had an average age of 20.38±1.03 when compared to male students 18,25±0.30. Regarding the perception of learning, there was a statistically significant difference in learning after the board game (p <0.05). Regarding the positive aspects of using the game, it was found that 23.5% reported that it helps to fix the content, 19.6% that provides integration between students, 18.9% that facilitates learning anatomy, 15.5% that raises students self‐esteem and 15.16% promotes better interpersonal relationships. The parameters of Reasoning, Safety, Learning, Evaluation Effectiveness, Speed and Cognition were statistically significant when compared to traditional teaching. It was found that the board game anatomy of the endocrine system is an important tool in the construction of anatomical knowledge, promoting dynamic, playful and meaningful learning, when compared with the traditional methodology.

Support or Funding Information

CNPQ and FUNCAP
Manifestation of Anxiety in Gait
Authors: Stark Maggie , Yager Chelsea , Taladay Phylicia , Teymouri Seema , Alvarez Julieth , Vogel-Rosbrook Christina , Martin Rebecca , Boolani Ali
Published: April 2020

In the United States, approximately 18% of adults are affected by anxiety. Nearly $42 billion are spent annually in the U.S. toward alleviating this mood disorder. However, only 10% of those with anxiety are receiving appropriate and effective treatment due to lack of identification. The purpose of this study was to identify gait patterns that are correlated with anxiety to improve probability of detection of this mood disorder. Past studies have examined gait and anxiety in Parkinson’s disease or in healthy individuals with induced anxiety. Our study tries to determine identifiable gait patterns that could signify the presence of increased anxiety in those without a diagnosed mood disorder. Participants (N=133, Males=50, Females=83, Age=25.8±7.96, BMI=24.8±3.8) ages 18 to 69 completed a Profile of Mood Survey (POMS) to assess current feelings of anxiety and completed a two‐minute walk around a 6 m track. Gait data was collected utilizing APDM mobility monitors. A backwards linear regression model was utilized to identify gait correlates of anxiety. The model predicted 13.1% of variance (R2=0.215, F= 2.553, p<.009) associated with anxiety. Our results showed that increased feelings of anxiety were associated with an increase in lateral anticipatory postural adjustment, lateral neck movement, toe‐out angle, transverse range of motion in the back, turn angles, and number of steps in a turn. Anxiety was also associated with decreased neck rotation, asymmetries in mid‐swing elevation, step variability, and sagittal range of motion in the back. Our results indicate that as feelings of anxiety increase in intensity, subjects present with a more narrow base of support, which may cause the increase in lateral movements during gait. Identifying gait patterns associated with anxiety mood disorders may lead to improved identification of individuals that may benefit from preventative and/or early intervention care plans. Future research should focus on whether increased anxiety is associated with a greater input from higher brain areas that overpower the rhythmic neural output of learned central pattern generators of gait. Additionally, caution should be used to ensure that multiple diagnoses are considered in the presentation of distinct characteristics of gait patterns.
Lysosomal PPT1‐insufficiency is a common pathogenic link between INCL and JNCL
Authors: Appu Abhilash Puthuvelvippel , Bagh Maria , Sadhukhan Tamal , Mondal Avisek , Mukherjee Anil B.
Published: April 2020

Neuronal Ceroid Lipofuscinoses (NCLs), also known as Batten disease, constitute a group of the most common inherited childhood neurodegenerative lysosomal storage disorders (LSDs). Even though mutations in >13 different genes cause various NCLs, at the cellular level, all the NCLs characteristically accumulate autofluorescent material (ceroid) in lysosomes and clinically, develop seizures, visual failure and a progressive decline in cognitive and motor functions. These findings suggest shared pathogenic mechanism(s) in these diseases. While inactivating mutations in the CLN1 gene, encoding lysosomal PPT1, cause infantile NCL (INCL), mutations in the CLN3 gene encoding a lysosomal membrane protein of unknown function underlie juvenile NCL (JNCL). PPT1 catalyzes depalmitoylation of S‐acylated proteins, required for their degradation by lysosomal hydrolases. Thus, PPT1‐deficiency causes lysosomal accumulation of S‐acylated proteins (ceroid) leading to INCL. The Cln1−/− and Cln3−/− mice are reliable animal models of INCL and JNCL, respectively. Here we report for the first time that Cln3‐protein requires S‐palmitoylation for its endosomal transport to the lysosomal membrane. Moreover, we found that in the brain of Cln3−/− mice, lysosomal PPT1‐protein as well as Ppt1‐enzymatic activity are significantly lower compared with lysosomes derived from their age‐ and sex‐matched WT counterparts. Unexpectedly, we found that the lysosomes from Cln3−/− mice accumulate significantly higher levels of S‐palmitoylated proteins, characteristically found in those from Cln1−/− mice. Previously, it has been reported that CLN3 mutations suppress the exit of cation‐independent mannose 6‐phosphate receptor (CI‐M6PR) from trans‐Golgi network (TGN), essential for its endosomal trafficking to the late endosome/lysosome. Since CI‐M6PR transports soluble proteins including Ppt1 to the lysosome, we hypothesized that Cln3 mutations may adversely affect the transport of PPT1 to the lysosome causing Ppt1‐insufficiency. Biochemical and confocal microscopic analyses of brain tissues from Cln3−/− mice revealed significantly higher levels of Ppt1 and CI‐M6PR in the Golgi compared to those of their WT counterparts. Collectively, our results demonstrate that lysosomal Ppt1‐insufficiency is a common pathogenic link between INCL and JNCL and suggest that a recently identified Ppt1‐mimetic, neuroprotective small molecule, NtBuHA, may have therapeutic implications for both INCL and JNCL.

Support or Funding Information

Intramural Research Program of the Eunice Kennedy Shriver NICHD, NIH
Expression and Immunolocalization of Xenobiotic Transporters at the Human Blood‐Testis Barrier
Authors: Hau Raymond Kin , Wright Stephen H. , Cherrington Nathan J.
Published: April 2020

The blood‐testis barrier (BTB) is formed by basal tight junctions between adjacent Sertoli cells of the seminiferous tubules and functions as a physical barrier to protect developing germ cells in the adluminal compartment from reproductive toxicants. Multiple xenobiotics are known to cross into the male genital tract (MGT) including male contraceptives, cancer chemotherapeutics, and environmental toxicants, although the mechanism(s) for their movement into the MGT is(are) largely unknown. Previous studies showed that several ATP‐binding cassette (ABC) and solute carrier (SLC) superfamily transporters are expressed in the testes and could potentially facilitate the disposition of xenobiotics into the MGT through concurrent uptake and efflux mechanisms. This study aimed to determine the expression and localization of xenobiotic transporters that may facilitate transepithelial transport across the human BTB. We used immunofluorescence staining to determine the cellular localization of these transporters in the several cell types found in human testes. OAT1, OAT2, OCTN1, and MRP3 were primarily localized to the basal membrane of Sertoli cells, whereas OCTN2, MRP6, and MRP7 were localized to both the basal membrane of Sertoli cells and the peritubular myoid cells that surround the basal membrane of Sertoli cells. Interestingly, CNT2 was localized to the lateral membranes of Sertoli cells and the peritubular myoid cells. OCT1, OCT2, and OCT3 were localized to the peritubular myoid cells and the Leydig cells, although there was moderate staining of spermatogenic cell membranes. However, definitive staining of CNT1, OAT3, MATE1, MATE2, OATP1A2, OATP1B1, OATP1B3, OATP1C1, OATP2A1, OATP2B1, OATP3A1, OATP4A1, and OATP6A1 was not observed due to non‐specific antibody staining or an absence of staining. The localization of CNT2, OAT1, OAT2, OCT1, OCT2, OCT3, OCTN1, OCTN2, MRP3, MRP6, and MRP7 suggests these transporters may contribute to the mechanism(s) by which xenobiotics can cross the BTB. Taken together, the localization of these pharmacologically relevant transporters provides insight into the selectivity of drug disposition across the BTB and may be useful in developing tools to understand and overcome the pharmacokinetic and pharmacodynamic difficulties presented by the BTB.

Support or Funding Information

5R01GM123643‐045R01GM129777‐02

P30CA023074
Retinoic Acid Receptor α Signaling Protects Against Diet‐induced Hepatosteatosis and Obesity in Mice
Authors: Bawa Fathima Nafrisha Cassim , Xu Yanyong , Zhu Yingdong , Chen Shaoru , Gopoju Raja , Zhang Yanqiao
Published: April 2020

Nonalcoholic fatty liver disease (NAFLD) is often characterized by accumulation of lipids in the liver. It presents a pathological spectrum of changes from simple steatosis to steatohepatitis. It is also often associated with obesity and insulin resistance. Since liver is the main organ involved in retinoid signaling, the impairment of this pathway is shown to increase liver fibrosis with the activation of stellate cells which stores retinoids in liver. However, the role of the retinoid signaling in NAFLD and obesity is not fully understood. Some studies have shown that retinoic acid treatment can ameliorate insulin resistance and obesity. We hypothesize that Retinoic Acid Receptor α (RARα) in hepatocytes play a significant role in mediating retinoid signaling to protect against NAFLD and obesity. To address this hypothesis, we conducted in vivo studies using liver‐specific Rarα knockout mice. Rarα floxed mice were injected with AAV8‐TBG‐Cre or AAV8‐TBG‐Null to generate liver‐specific Rarα knockout mice (L‐Rarα−/−) mice and the control mice (Rarαfl/fl). These mice were gavaged with either vehicle or all‐trans retinoic acid (AtRA; 15mg/kg/day) along with high fat/high cholesterol/high fructose (HFCF) diet for 16 weeks. The data showed that AtRA treatment reduced body fat content and increased energy expenditure in the control mice but not in L‐Rarα−/− mice. In addition, AtRA reduced hepatic triglyceride (TG) levels in both the control mice and L‐Rarα−/− mice with a greater extent in reduction of hepatic TG levels in the control mice. Analysis of hepatic gene expression by qRT‐PCR showed that AtRA reduced hepatic levels of genes involved in lipogenesis and fatty acid uptake (Pparγ, Cd36 and L‐Fabp) or inflammation (Tnf‐α) in the control mice, but not in L‐RARα−/− mice. Similar observations were collected in a different study where control mice and L‐Rara−/− mice were fed an HFCF diet for 20 weeks, followed by 10 weeks of treatment with either vehicle or AtRA. Our data demonstrate that the retinoid signaling protects against the development of hepatosteatosis and obesity in a Rarα‐dependent manner.

Support or Funding Information

Support or funding information: R01DK118805 and R01HL103227
Brazilian Berry Extract, High‐Fat Diet and Prostate Cancer: In Vitro and In Vivo Evaluation of Inflammation and Extracellular Matrix
Authors: Cagnon Valéria H. , Lima Ellen Nogueira , Lamas Celina de Almeida , Baseggio Andressa Mara , Kido Larissa Akemi , Rosseto Isabela Urra , Santos Felipe R. , Montico Fabio , Maróstica Mario Roberto
Published: April 2020

The high incidence of prostatic cancer (PCa) has encouraged studies in prostatic cellular biology all over the world. In addition, Brazilian berries, such as Myrciaria jaboticaba (Vell.) Berg, show a high polyphenol concentration in the peel, characterizing antioxidative and anti‐inflammatory properties. The aim herein was to analyze the anti‐inflammatory and remodeling tissue effects of the Jaboticaba peel extract (JPE), a patented product, in early prostatic adenocarcinoma development associated with high‐fat diet (HFD) intake, as well as to investigate the JPE role in human PCa and LNCaP cell lines. TRAMP mice from control groups received standard diet or HFD and the treated groups followed the same diet protocol plus JPE (5.8g/Kg/body weight during 5 days per week for 2 months). The dorsolateral prostate was collected for immunohistochemistry and Western Blotting analyses. PC‐3 and LNCaP cells were treated with different doses of JPE (125, 250, 500 ug/mL) and used for cellular viability assay and Western Blotting analyses. The results showed that the HFD intake led to an MMP‐9 increase and a Laminin protein level decrease, worsening the PCa profile in this transgenic model. Conversely, JPE led to decreased MMP‐9 and also TGF‐beta after HFD intake. Laminin level increased in both groups treated with JPE. Dose and time dependent JPE exposure reduced cellular viability, especially, in androgen‐responsive LNCaP cells. In addition, JPE induced NfkB and iNOS reduction not only in LNCaP but also in PC‐3 cell lines, suggesting that the growth inhibitory effect of JPE in PCa may be associated with its anti‐inflammatory role. To conclude, the tissue damage caused by HFD led to the deregulation of the extracellular matrix elements, which intensified cancer progression, compromising the epithelial‐stromal interaction in the dorsolateral prostate. However, JPE was beneficial for the dynamic interaction between the epithelium and stroma in vivo, and altered cancer cell growth by means of anti‐inflammatory pathway, pointing towards a possible adjuvant therapy for PCa delay, highlighting the natural compounds as an important strategy in prostatic disorders.

Support or Funding Information

Sao Paulo Research Foundation/FAPESP (2018 04579‐7). And Fund for Support to Teaching, Research and Outreach Activities FAEPEX/UNICAMP (2080/20)
Photoreceptor Cell Development Requires CDHR1a Protein Stability Controlled by Siah E3 Ubiquitin Ligase During Zebrafish Eye Development
Authors: Piedade Warlen Pereira , Titialii Kayla , Morris Ann , Famulski Jakub
Published: April 2020

E3 ubiquitin ligases mediate orderly and precise targeting of protein degradation to maintain biological homeostasis and coordinate proper development. Our lab recently discovered that Siah family of E3 ligases plays a role in early ocular morphogenesis, in particular fusion of the optic fissure. Interestingly, Siah ligases are also expressed during photoreceptor development and are predicted to target CDHR1a, a cadherin superfamily of calcium‐dependent cell adhesion molecules and photoreceptor‐specific cadherin. Mutations in this cadherin are associated with inherited retinal dystrophies, such as cone‐rod dystrophy. Using whole mount in situ hybridization and Immuno‐histochemistry we detected Siah gene expression and protein localization in the outer nuclear layer (ONL) and in the retinal ganglion cell layer starting at 3 days‐post fertilization (dpf) until 7dpf. CDHR1a expression and protein localization overlaps with both Siah genes in the outer segment of the retina from 3 until 7dpf. We therefore hypothesized that Siah regulates CDHR1a during photoreceptor development. To test our hypothesis, I first confirmed siah‐mediated targeting of CDHR1a for degradation using a mammalian cell culture, co‐immunoprecipitation followed by western blotting analysis. Second, I created two transgenic zebrafish lines that express Siah1 or a dominant negative (Siah1DRING) under the control of the heat shock promoter. Using the heat shock line to overexpress Siah1 at 2dpf and 2.5dpf, we observed a decrease in the number of developing rods and cones in the retina at 3dpf. The number of retinal ganglion cells, amacrine, bipolar and horizontal cells, however, did not change. In addition, there was a significant reduction of proliferating (EdU+ and PH3+ cells in ONL) cells in the retina. Taken together, our results suggest that Siah ubiquitin ligases may control CDHR1a stability and therefore regulate the photoreceptor cell proliferation and differentiation.

Support or Funding Information

This work was supported by the Brazilian National Council for Scientific and Technological Development (CNPq) under grant number 202970/2014‐0.
Four‐week Ketogenic Diet Improves Aerobic Metabolic Capacity in Extensor Digitrum Longus Muscle of Sedentary Male Rats
Authors: Ogura Yuji , Yoshihara Toshinori , Kurosaka Mitsutoshi , Kakigi Ryo , Fujiwara Sei-etsu , Kakehashi Chiaki , Higashida Kazuhiko , Funabashi Toshiya
Published: April 2020

Ketogenic diet is an extremely high‐fat diet with low or absence of carbohydrate in the total nutrient. Thus, this diet causes ketone body production from the liver and shifts energy sources from carbohydrate to fat. Whereas previous studies have suggested that ketogenic diet ameliorates brain disorder such as drug‐resistant epilepsy, its influence on peripheral tissue is unclear. Skeletal muscle is an important organ for locomotion and metabolism. Recent studies have suggested that the ketogenic diet changes the energy metabolic properties in the skeletal muscle as well; however, the results are contradictory. Moreover, it remains unknown whether the ketogenic diet affects the muscle contractile function. Therefore, this study aimed to clarify the effect of ketogenic diet on muscle performance and biochemical adaptation in rat skeletal muscle. This study was approved by Institute for Animal Experimentation at St. Marianna University School of Medicine. Male Wistar rats (8 weeks of age) were assigned to either control group (CON) or ketogenic diet group (KD). CON and KD were fed for 4 weeks either control diet (10% fat, 10% protein, 80% carbohydrate) or ketogenic diet (90% fat, 10% protein, 0% carbohydrate), respectively. Body mass was recorded once a week. After the 4‐week feeding period, the animals were anesthetized, and the blood sample was collected to analyze blood ketone concentration. The right extensor digitrum longus (EDL) muscle was used to evaluate the twitch force, tetanic force, and fatigue ex vivo. The left EDL was homogenized and used to analyze the myosin heavy chain composition, the protein expression of metabolic enzymes, and the citrate synthase activity. Body mass was lower in KD than CON at third and fourth week of the feeding period. Blood ketone concentration in KD was significantly higher than CON. There was no statistical difference in the twitch and tetanic forces as well as muscle fatigue between CON and KD. However, the citrate synthase activity in KD was significantly higher than CON. The protein expression of citrate synthase, succinate dehydrogenase, cytochrome c oxidase subunit 4, and 3‐hydroxyacyl‐CoA dehydrogenase in KD was significantly higher than CON. Moreover, there was a shift of myosin heavy chain from type IIb to IIx in KD. Therefore, we conclude that 4‐week ketogenic diet improves skeletal muscle aerobic capacity without obstructing muscle contractile function in the sedentary male rats.

Support or Funding Information

This work was supported by MEXT/JSPS KAKENHI Grant Numbers JP24650334, JP18K17767.
The Application of 3D Printing and Nutrient/Biomaterial Microhabitats for In Situ Enrichment of Microbial Cultures
Authors: Andersen Morten , Moreira-Grez Benjamin , Andersen Magdalena Julia , Whiteley Andrew
Published: April 2020

The objective of this study was to develop a new tool for selectively capturing new microorganisms from the environment. High‐throughput environmental sequencing has revealed a diversity much greater than that which we have brought into culture. Even so, the microorganisms that have been successfully cultured have yielded a wealth of pharmaceuticals, enzymes and living cultures for medical, agricultural and industrial use. Bringing new microorganisms into culture thus has great value.

A MicroCaptureArray for high‐throughput in situ enrichment of new species from the environment using metabolic cues was created using 3D printing. The system is based on repurposing biomaterials developed for medical implants and controlled release devices
[1],
[2]. The MicroCaptureArray was tested in a terrestrial (soil) and an aquatic (pond) environment for 9 and 7 days, respectively. DNA was then isolated from the array and subjected to 16S sequencing with Qiime2 data analysis.

The MicroCaptureArray attracted, captured and enriched different species, many of which were undetectable in the background community using 16S sequencing. The different metabolic cues resulted in the enrichment of different microorganisms. The system also enriched potentially new species, in several samples more then 25% of the captured microorganisms were unassignable beyond the Kingdom of Bacteria or the Phylum of Proteobacteria, in constrast, unassignable bacteria accounted for less than 0.3% of the background samples.

In conclusion, the MicroCaptureArray shows that metabolic cues may be used for in situ baiting and enrichment culture of bacteria. The tool may allow researchers to detect rare microorganisms that are present but undetectable in the environment that may nonetheless play a major role when certain nutrients are introduced. It may also allow selective attraction and enrichment of environmental bacteria that are capable of metabolizing particular substances. This may allow targeted discovery of entirely new species with desirable properties.

Support or Funding Information

The project was supported by the Villum Foundation (grant number 17627)
Engineering Breast Cancer Mammospheres Using Vibration
Authors: Jolly Mehma K. , Sengupta Shiladitya , Dai Hongqing , Jang Haelin
Published: April 2020

Cancer organoids are increasingly being used as choice model for studying cancer biology. However, creating uniform cancer organoids is a challenge. Here we describe an easy method to engineer uniform cancer organoids. In our experiment, we generated calcium‐alginate drops to encapsulate cells and use as 3D tumor spheroid models. We used a low‐cost, oil‐free, simple and efficient method for making uniform drops so our results could be accurate. We added a periodic vibration to a continuous jet of alginate solution mixed with breast cancer cells by using a mini rotation motor and we let the drops fall into a calcium bath. The job of the vibration is to cut the jet into drops of the same size, but there were other parameters that changed the results. To ensure uniformity, we adjusted the following parameters: the nozzle tip diameter, the material of the nozzle, the vibration frequency, and the velocity of the syringe pump that produced the jet. After finding the best combinations of all the parameters, we were able to produce uniform drops of different sizes. We found that the breast cancer cells were encapsulated so the droplets act as a more realistic and efficient way to model tumors. These tumor models can be used to test drug penetration and find new anticancer treatments. The models can also be modified to simulate cancer size and concentration, and measure differences in drug penetration efficacy as a result.
Computational Analysis of Wild‐type and Selenium‐Incorporated Down‐regulated Proteins in E. coli
Authors: Hill Austin Reece , Cashman Derek J. , Boles Jeffrey Oakley
Published: April 2020

The addition of heavy metal amino acid analogs has long been a standard augmentation of X‐ray crystallography to solve the phase problem associated with smaller atoms such as carbon, nitrogen, and sulfur. Unfortunately, the addition of such analogs, like selenomethionine (SeMet), brings about unprecedented negative implications concerning the growth of the exposed cell cultures. The negative impact of growth is due to the cytotoxic effects of selenium‐incorporation, either partial or complete, as seen by a proteomic analysis performed by Broderick and Boles in 2013. It is apparent that there are a marked number of completely down‐regulated key enzymes in these exposed cell cultures. Many aspects of the down‐regulation cannot be experimentally tested; thus, computational methodology can be utilized to provide unique insight into the cellular processes and the cytotoxic effects of selenium on proteins in vivo. An AMBER14 force field has been modified to properly model selenium atoms in the protein structure. This modification allows for the analysis of protein stability and protein/substrate interactions of the mutant variant down‐regulated enzymes.
FLG LOSS OF FUNCTION MUTATIONS IN PUERTO RICO: ASSOCIATION OF GENETIC ANCESTRY WITH ATOPIC DERMATITIS AND ICHTHYOSIS VULGARIS
Authors: Aponte Edwin Gilberto Ramirez , Cruzado Juan Carlos Martinez
Published: April 2020

FLG mutations R501X, R2447X and S761fs have been associated with populations in Europe, but they are also present in admixed populations with European ancestry. The polymorphism Q3818X, in the 1000 Genomes Project, appears to be only present in Puerto Rico and it does not appear to have a severe effect on the structure of the filaggrin protein. Because it has never been found outside of the island, it has not been studied nor related to IV or AD. Population genetic data for these mutations outside of the 1000 Genomes Project is unknown for the island and this study attempts to be the first genetic study focused on establishing their allele frequencies in the general population, and in the affected population. A TaqMan genotyping using qPCR will be employed to establish the frequency of all SNPs. A total of 684 samples from 47 municipalities are going to be genotyped for all mutations. For the affected population, the number of samples required using a correlation of 0.2, a nominal power of 0.90 and α of 0.05 will be 211 with AD and 211 with IV. We already genotyped 736 samples from 47 municipalities for R501X and found a higher allele frequency than the one previously reported; and higher frequency in the South‐Center Region of the island that could be related to the European ancestry distribution. A total of 292 samples from 29 municipalities were genotyped for R2447X and we found a lower allelic frequency in our study and a regionalization in the East Region. We expect to do the same for the other genetic variants and genotyped all of them in the affected population. Data and educational information will be shared with the public, dermatologist and physician in the island to increase the diagnosis of AD and IV in the island.
Inhibitory effect of Sihocheonggan‐tang on 2,4‐dinitrochlorobenzene‐induced atopic dermatitis in BALB/c mice
Authors: Chun Jaemoo , Park Seonghwan , Jung Jeeyoun
Published: April 2020

Atopic dermatitis (AD) is a chronic inflammatory skin disease characterized by complex immune dysregulation. Although the prevalence of AD has increased steadily in recent years, therapeutic treatments remain limited. In this study, we investigated the effects of Sihocheonggan‐tang (SHCG) on AD induced by 2,4‐dinitrochlorobenzene (DNCB) in BALB/c mice. Repeated application of DNCB was performed on the dorsal skin of BALB/c mice, and SHCG was orally administered daily for 2 weeks. We found that SHCG administration alleviated DNCB‐induced AD symptoms such as skin lesion and clinical skin severity score. Histopathological analysis indicated that SHCG significantly reduced the epithelial skin thickness and decreased the expression of thymic stromal lymphopoietin (TSLP), intercellular adhesion molecule‐1 (ICAM‐1), and mast cell infiltration in skin. Also, SHCG administration alleviated the increased immunoglobulin E (IgE) and Th2 cytokines such as IL‐4, TNF‐α, and IL‐6, induced by DNCB in BALB/c mice, suggesting that SHCG suppresses AD via the regulation of Th2 immune responses. Furthermore, we investigated the gut microbiota to clarify the association between AD and composition of gut microbiota. SHCG administration attenuated the abundance of Deferribacteres, which was increased in DNBC‐induced AD mice. Principal coordinate analysis (PCoA) revealed a separation of SHCG group from AD group at the species level. These results suggest that SHCG may be a promising therapeutic candidate for the treatment of AD by modulating gut microbiota composition.
GPR55 Regulates Progression of Atherosclerosis by Monocyte Adhesion to Vascular Endothelium
Authors: Lee Seung-Jin , Im Dong-Soon
Published: April 2020

G‐protein coupled receptor 55 (GPR55) is an orphan G‐protein coupled receptor, which is participate in the progression of atherosclerosis. however, the underlying mechanisms are unclear. Thus, this study investigated the molecular mechanisms responsible for monocyte adhesion to the vascular endothelium, which is a key early event in macrophage foam cell formation. In in vitro study, the pharmacological GPR55 agonist, O‐1602‐induced monocyte adhesion to the endothelial cells and adhesion molecule, Mac‐1 expression were significantly attenuated by pretreatment of GPR55 antagonist, CID 164002 in monocytes. In addition, in promoter activity analysis and chromatin immunoprecipitation assays to identify transcription factors involved in O‐1602‐induced Mac‐1 expression, both AP‐1 and NF‐κB played central roles to increase Mac‐1 transcription in O‐1602‐treated monocytes. Moreover, O‐1602‐induced activations of Gq/11 proteins, CaMKK, and AKT were found to be signaling components of the monocyte adhesion to the endothelial cells and Mac‐1 expression pathway. In in vivo study, a greater number of atherosclerotic plaques formed in ApoE(−/−) mice fed a high‐fat diet than in CID 164002 pretreated ApoE(−/−) mice, indicating a pivotal role for GPR55 in preventing vascular inflammation. Based on these results, GPR55‐mediated expression of Mac‐1 in monocytes plays a pivotal role on monocyte adhesion to vascular endothelium, leading to increased foam cell formation in the development of atherosclerosis.