bluuebunny/crossref_metadata_2025 huggingface dataset:
Autonomic dysfunction precedes cardiometabolic alterations in a metabolic syndrome model: impact of aerobic exercise training
Authors: Dias Danielle , Bernardes Nathalia , Conti Filipe Fernandes , Brito Janaina Oliveira , Irigoyen Maria Claudia , De Angelis Kátia
Published: November 2023

The aim was to evaluate the effects of exercise training (T) on the time course of cardiometabolic alterations in hypertensive rats (SHR) submitted to fructose overload. Wistar and SHR rats were divided: Control (C), Hypertension (H), Hypertensive + Fructose (HF) and Hypertensive Fructose + Exercise Training (HFT). The fructose overload (100g / L) was initiated 30 days after weaning of the animals. The T was performed on a treadmill (1H / d, 5d / week). The evaluations were performed at 37, 45, 60 and 90 days of life of animals. Blood pressure (BP) and heart rate (HR) were directly recorded through an acquisition system. Cardiovascular autonomic modulation was evaluated in the time and frequency domain. The H and HF groups showed an increase in VAR‐SAP compared to C group at all times. The consumption of fructose induced a further rise of the VAR‐SAP at 45 and 90 days vs. 37 days, which was not observed with the T. The H group showed an increase in LF‐SAP (vascular sympathetic) at 60 and 90 days compared to C group. However, the HF group showed an increase in VAR‐SAP at 45, 60 and 90 days compared to C group and their values in 37 days, with an additional increase in 90 days (40%) vs. H group. The H group showed a decrease in HF‐PI (cardiac parasympathetic) in 37 days and the HF group at 37 and 90 days compared to C group. The HFT group had higher RMSSD and VAR‐PI at 37 and 90 days compared to C group. The spontaneous baroreflex sensitivity was lower in H and HF groups in all times evaluated and the T increased this variable in 60 (79%) and 90 days (72%) vs. HF group. The H and HF groups showed an increase of BP in the all evaluated times compared to C group. (165±3; 179±3 vs. 114±1 mmHg) The fructose induced an additional increase in BP at 90 days and T reduced these values. The HFT group showed a HR reduction in 60 (11%) and 90 (7%) days vs. HF group. There were no significant changes in blood triglycerides and glucose in all evaluated times. The cardiovascular autonomic dysfunction seems to precede hemodynamic and metabolic changes in this experimental model of metabolic syndrome. Moreover, exercise training was effective in attenuate the time course of autonomic and hemodynamic dysfunctions.

Support or Funding Information

Financial Support: FAPESP (2015/11223‐6); CNPq (457200/2014‐6); CAPES (PROSUP).
Antioxidants Attenuate the Exercise Induced Resetting of the Arterial Baroreflex in Healthy Human Subjects: Implications for Exercise Induced Hypertension
Authors: Jouett Noah , Moralez Gilbert , White Daniel Wade , Raven Peter Bernard
Published: November 2023

Patients with Exercise‐induced‐Hypertension (EiHT) exhibit exaggerated increases in arterial pressure at the onset of exercise which may prevent EiHT patients from participating in exercise training programs. EiHT is thought to occur due to dysregulated resetting of the arterial baroreflex (ABR). Prior studies in animal models demonstrate that reactive oxygen species (ROS) generated in the brainstem scavenge the sympathoinhibitory function of central Nitric Oxide (NO) and, thereby enable ABR resetting of the operating point (OP) pressure and hypertension. We tested the hypothesis that a centrally and peripherally active antioxidant cocktail (CT; composed of Vitamin E and C with Co‐Q10) will attenuate the exercise induced resetting of the ABR's centering point (CP) and OP pressures compared to the same exercise intensity performed with a vehicle placebo (PL). Seven healthy human subjects were recruited and performed 700 back‐supported semi‐recumbent dynamic leg exercise at moderate (HR at 120 beats per minute: e120) and heavy (HR at 150 beats per minute: e150) intensities. Mean arterial pressure (MAP) was continuously recorded using photoplethysmography at the finger, while HR was recorded via a three lead electrocardiogram (ECG). On experimental day 1, subjects were either given the CT or PL 1 hr. (time of peak plasma concentrations) prior to the start of exercise. On a separate experiment day 2, the subjects repeated the same exercise intensity protocol with the other test article (CT or PL) in a randomized repeated measures design. During exercise with the PL ingestion, the CP of the ABR was reset to higher MAPs from rest to e120 (100 ± 3 mm Hg to 121 ± 3 mm Hg, P<0.02) but not e150 (113 ± 3 mm Hg, P=0.15). The absence of resetting at the higher work intensity was likely due to cardiovascular drift (decreasing MAP). Ingestion of the CT prior to the exercise protocols prevented the increase of the CP to higher MAPs from rest to e120 and e150 (rest: 97 ± 3 mm Hg, e120: 106 ± 3 mm Hg, e150: 106 ± 3 mm Hg, P >0.21). Furthermore, the OP‐pressure of the ABR was attenuated with CT ingestion compared to PL at e120 (placebo e120: 116 ± 0.8 mm Hg, CT e120: 111 ± 0.8 mm Hg, P = 0.04). These data: (a) confirm that centrally derived ROS contribute to exercise induced ABR resetting; and (b) indicate that EiHT could be treated by ingestion of an anti‐oxidant cocktail prior to the start of exercise
Modulation of Methylated Arginine Metabolism to Target Exercise‐Induced Fatigue in Duchenne Muscular Dystrophy
Authors: Garbincius Joanne F. , Merz Lauren E. , Cuttitta Ashley J. , Bayne Kaitlynn V. , Schrade Sara , Armstead Emily A. , Whitesall Steven E. , D'Alecy Louis G. , Michele Daniel E.
Published: November 2023

Duchenne muscular dystrophy (DMD) is an X‐linked disease caused by mutations in dystrophin and characterized by muscle degeneration, cardiomyopathy, and impaired muscle nitric oxide (NO) production that leads to misregulated muscle blood flow and excessive post‐exercise fatigue. Data from our laboratory show that serum levels of the NO synthase inhibitor asymmetric dimethylarginine (ADMA) are elevated in dystrophin‐deficient mdx mice and after acute muscle injury, and that infusion of ADMA is sufficient to impair exercise performance in wild‐type mice. Therefore, we hypothesized that excessive circulating ADMA impairs NO production and exaggerates exercise‐induced fatigue in DMD. To determine the impact of ADMA on fatigue resistance in this disease, female mdx mice were crossed to wild‐type males expressing a transgene encoding human dimethylarginine dimethylaminohydrolase 1 (DDAH), an enzyme that degrades ADMA. Voluntary ambulatory activity of transgenic (TG) and non‐transgenic (NTG) offspring was measured in open field assays prior to and following acute bouts of treadmill running to assess exercise‐induced fatigue. DDAH expression lowered serum ADMA concentration in TG offspring, but failed to attenuate the post‐exercise decrease in voluntary activity in mdx males. However, transgene expression resulted in a significant, 20% attenuation of the decrease in voluntary activity following exercise in female mdx carriers. The heart weight/tibia length ratio was significantly reduced in TG versus NTG carriers, although examination of the gastrocnemius and soleus muscles revealed no difference between genotypes in histological markers of dystrophy or ischemic muscle injury, suggesting that improved fatigue resistance was due to cardiac‐specific effects of DDAH expression. We conclude that ADMA contributes to excessive exercise‐induced fatigue in dystrophin heterozygous females, possibly by promoting pathological cardiac remodeling and impairing heart performance. These findings emphasize the importance of NO signaling to cardiac function in dystrophinopathies and reveal a new therapeutic target to mitigate the risk of cardiomyopathy in DMD carriers.

Support or Funding Information

Supported by the Muscular Dystrophy Association, the University of Michigan CVRE Training Program, and NIH T32 GM‐08322.
3T3‐L1 Adipocytes Take Up Norepinephrine: No Canonical Norepinephrine Transporters Found
Authors: Ismail Alex , Ayala‐Lopez Nadia , Burnett Robert , Watts Stephanie
Published: November 2023

Rat perivascular adipose tissue (PVAT) and rat adipocytes contain the organic cation transporter 3 (OCT3), a transporter of norepinephrine (NE). This is physiologically relevant because PVAT reduces vasoconstriction through the uptake of NE. We hypothesized that an accepted model of adipocytes, 3T3‐L1 cells, take up NE through OCT3 or a similar catecholamine transporter. Mouse 3T3‐L1 fibroblasts were induced into adipocytes by adding 0.5 mM IBMX and 0.25 mM dexamethasone for 48 hours. AdipoRedTM assay and Oil Red O staining confirm that induction was successful (15 times more fluorescent than preadipocytes and 95% of cells exhibit lipid droplet staining, respectively) on the 10th day after the induction. NE was added to 3T3‐L1 adipocytes at 0, 1, 10, and 100 mM concentrations in physiological saline solution (PSS) with NE metabolism inhibitors [monoamine oxidase, pargyline (10−5 M), catecholamine‐O‐methyltransferase, Ro 41‐0960 (10−6 M), semicarbazide‐sensitive amine oxidase, semicarbazide hydrochloride (10−3 M)] for 30 minutes at 37 °C. Cells were washed with PSS, collected in tissue buffer, and analyzed for NE content using HPLC. There was an increase in NE, normalized for protein content, from vehicle to 10 mM and 100 mM (0.09 ± 0.01 pg/mg, 5.97 ± 1.48 pg/mg, 18.89 ± 3.37 pg/mg, respectively, p < 0.05, N=6). PCR was performed on cDNA isolated from 3T3‐L1 adipocytes to test for known NE transporters: norepinephrine transporter (NET), OCT3, serotonin transporter (SERT), dopamine transporter (DAT), vesicular monoamine transporter 2 (VMAT2), and plasma membrane monoamine transporter (PMAT), using beta‐actin as the reference gene. OCT3, SERT, DAT, and VMAT2 did not reach the detectable threshold after 40 amplification cycles (CT value > 40, N=4). NET and PMAT had 1 sample out of 4 with a detected CT value (37.45 ± 0.15 and 37.41 ± 0.05, respectively). The positive controls had detectable amplification (heart: 26.94 ± 0.02 for OCT3, brain: 27.53 ± 0.10 for SERT, 27.90 ± 0.03 for DAT, and 26.97 ± 0.03 for PMAT, adrenal: 25.80 ± 0.15 for VMAT2, pons: 24.59 for NET). These data suggest that NE is entering 3T3‐L1 adipocytes via mechanisms other than the canonical transporters of NE. Learning how NE interacts with adipocytes, and therefore PVAT, is essential for studying the relation between vasoconstriction and obesity.

Support or Funding Information

NIH NHLBI P01HL70687

IPSTP 5 T32 GM 92715‐4

NIH NRSA F31 HL128035‐01
Effect of Vitamin D on Adipogenesis under Hypoxia
Authors: Mutt Shivaprakash Jagalur , Lehtonen Siri , Lehenkari Petri , Leppäluoto Juhani , Järvelin Marjo‐Riitta , Herzig Karl‐Heinz
Published: November 2023

Background

Obesity and pandemic vitamin D (vitD) deficiency both share similar pathological associations with adipose tissue chronic inflammation leading to well‐known conditions such as insulin resistance and hypertension. Understanding adipogenesis is of major relevance for human health, as increased adiposity leads to cellular hypoxia followed by inflammation, oxidative damage and adipocyte dysfunction resulting in metabolic diseases. We have shown earlier that vitD supplementation has significant beneficial effects on adipocyte inflammation and improved insulin sensitivity in vitD deficient mice. The modulation of adipogenesis under these circumstances might be beneficial in improving its metabolic functioning. Therefore, we hypothesize that vitD modulates adipogenesis in pathological conditions (hypoxia).

Methods

Human mesenchymal stem cells (hMSCs) were differentiated into adipocytes ± vitD [10−7 M] containing adipogenic induction and maintenance medium under either hypoxic (1% O2) or normoxic (21% O2) conditions. At the end of differentiation cycles the process was assessed by the oil red O staining of the adipocytes. Fully differentiated adipocytes were stimulated with lipopolysaccharide (LPS 10 ng/ml) ± vitD [10−7 M] for 24 hrs and cells and cell culture supernatants were collected from both hypoxic and normoxic basal ± vitD [10−7 M] as well LPS stimulated ± vitD [10−7 M]. The inflammatory marker interleukin‐6 (IL‐6) levels were measured by ELISA. RNA and cell lysate were prepared and analyzed for adipogenic, hypoxic and inflammation regulating proteins.

Results

The adipogenic differentiation of hMSCs under hypoxia was significantly inhibited in comparison to the normoxia. VitD [10−9 M to 10−7 M] significantly enhanced adipogenic differentiations dose dependently under both hypoxic and normoxic culture conditions. VitD [10−7 M] inhibited basal IL‐6 secretion by 83% in hypoxic and 66% in normoxic hMSCs differentiated adipocytes in comparison to their respective controls (p<0.05). In addition, vitD dose dependently reduced LPS induced IL‐6 secretion from hMSCs differentiated adipocytes under both hypoxic (10−7M:10−8M:10−7M; 66%:40%:19%) and normoxic (10−7M:10−8M:10−7M; 56%:44%:2%) condition in comparison with the LPS 10 ng/ml alone.

Conclusion

Our data demonstrates that vitD enhances the adipogensis of hMSCs under hypoxic culture condition; it inhibits basal IL‐6 release as well LPS induced IL‐6 release under both hypoxia and normoxia. This suggests that sufficient vitD levels would be able to modulate the hypoxia induced inflammation and adipogenic differentiation potential in obese subjects. These vitD actions might therefore partially mediate the beneficial effect on insulin sensitivity of adipose tissue and reduce the risk of known co‐morbidities of obesity.

Support or Funding Information

Supported in part by the Academy of Finland and DynaHEALTH EU No.633595
The Effect of Cholesterol on the Biophysical Inhibition of Pulmonary Surfactant by Albumin
Authors: Veldhuizen Ruud , Milos Scott , Ruehlicke Jake , Yamashita Cory
Published: November 2023

Pulmonary surfactant is a mixture of 85% phospholipids, 5–8% cholesterol and 5–8% surfactant proteins which forms a surface tension reducing lipid film at the air‐liquid interface of the alveoli. By reducing the surface tension to near 0mN/m values during compression (i.e. exhalation) surfactant maintains alveolar stability at low lung volumes and maintains proper lung compliance. The importance of surfactant is evident in Acute Respiratory Distress Syndrome (ARDS) in which the dysfunction of surfactant contributes to hypoxemia, regional areas of collapse and reduced lung compliance. Specifically, serum proteins, such as Albumin, leaking into the lung during ARDS have been shown to interfere with surfactant's ability to reach low surface tension. In addition, recent studies suggest that elevated cholesterol within surfactant also contribute to surfactant dysfunction in the setting of lung injury. Although the effects of cholesterol and serum proteins with surfactant have been studied independently, the interaction between the two has not previously been investigated. It was hypothesized that elevated levels of cholesterol within surfactant will make surfactant more susceptible to serum protein inhibition.

Methods

Bovine Lipid Extract Surfactant (BLES), a commercially available exogenous surfactant, was utilized and modified to generate samples with concentrations of 0, 2.5, 5 and 10% (w/w) cholesterol. Varying amounts of Bovine Serum Albumin (BSA) were subsequently added to each sample to attain concentrations of 0, 20, 30, 40 and 50 mg/ml. A constrained sessile drop surfactometer was used to determine the minimal achievable surface tensions during dynamic compression/expansion cycles.

Results

Samples with various levels of cholesterol, in the absence of BSA, reached minimum surface tension values of near 0mN/m during compression. At moderate levels of BSA (20 & 30mg/ml), minimum surface tensions were lower in surfactant samples containing cholesterol as compared to samples containing no cholesterol. Finally, at high BSA values, all surfactant samples had surface tension values that were higher than those at the lower BSA concentration but were not significantly different among the sample with different cholesterol concentrations.

Discussion

Contrary to the hypothesis, values of up to 10% cholesterol within surfactant do not appear to increase the susceptibility of surfactant to serum protein inhibition. In fact, the results suggest that cholesterol mitigates the effect of moderate levels of BSA on surfactant function. It is concluded that cholesterol within surfactant contributes to its resistance against inhibition by albumin.

Support or Funding Information

Canadian Institutes for Health Research.
Increased Alveolar Size Persists in a Rodent Model of Bronchopulmonary Dysplasia
Authors: Centanni Ryan , Haraldsdottir Kristin , Braun Rudolf , Barton Greg , Goss Kara , Eldridge Marlowe , Pegelow David
Published: November 2023

Introduction

Bronchopulmonary dysplasia (BPD) is a chronic lung disease associated with premature birth. Hallmark pathologic features of BPD include increased alveolar wall thickness and reduced pulmonary vascular cross sectional area. Measurement of mean linear intercept (MLI) is a tool used to quantify alveolar size, where a higher MLI is suggestive of reduced alveolar surface area. Previous studies have shown an increased MLI in lungs from a BPD rat model in early postnatal life. However, it is unknown if these pathological changes last into adulthood. The purpose of this study was to quantify alveolar size using the mean linear intercept (MLI) technique in lung tissue sections from 7 and 14 day and 1 year old control and BPD rats.

Methods

Sprague Dawley rat pups were placed in hyperoxic (HYP, 85% O2) or normoxic (NORM, 21% O2) chambers within 12 hours of birth for a total of 14 days, then were moved to room air housing. At 7 and 14 days and 1 year of age, rats (n=2 rats per time point per exposure) were sacrificed and lung tissue sections were placed on slides and stained with hematoxylin and eosin (H&E). 9 images from each lung cross section were collected and analyzed. Alveolar air space was determined using the MLI method where horizontal grids were placed on images and intersections with the tissue were counted. For each image, MLI was calculated using the following equation: Lm = number of intersections/(grid length (um) × number of lines). The 9 values were averaged, giving an MLI score for each animal. Two‐way ANOVA was used to determine the main effect for age (7, 14, 360 d), condition (NORM vs HYP), and the interaction between age and condition. Statistical significance was defined as p < 0.05.

Results

HYP rats have a significantly greater MLI (p=0.01) compared to NORM rats at all time points. At 7 days, HYP rats had a 43% increase in MLI compared to NORM. At 14 days, HYP rats had an 18.5% increase. At one year, HYP rats had a 47% increase.

Conclusion

Mean linear intercept is significantly increased a rat model of BPD, which persists well into adulthood. Enlargement of the distal airspaces likely plays a role in the diminished gas exchange observed in BPD.

Support or Funding Information

NHLBI, R01 HL115061‐03, Suppl (Eldridge) & NHLBI, R01 HL115061 (Eldridge)
Mesenchymal and Epithelial Specific Deletion of Rac1 Prevents Lung Morphogenesis
Authors: Danopoulos Soula , Krainock Michael R. , Al Alam Denise
Published: November 2023

Background

Epithelial‐mesenchymal interactions control specification, budding and branching in the developing lung. It is the spatio‐temporal expression of various signaling molecules in the different tissue compartments that drive the patterning of the lung. Our previous work demonstrated that the inhibition of Rac1 in mouse lung explants result in impaired branching morphogenesis and vasculogenesis. This role was primarily due to impaired mesenchymal cell proliferation and differentiation following Rac1 inhibtion. Therefore, we sought to determine the effect of epithelial and mesenchymal Rac1 deletion in vivo on lung development during the pseudoglandular and saccular stages.

Methods

To assess the effect of loss of epithelial or mesenchymal Rac1 on lung development in vivo, Rac1f/f animals were used to conditionally inactivate Rac1 using a Shh‐Cre or Dermo1‐Cre driver respectively. The lungs were then harvested from E13.5, E14.5 and E18.5 mutant (Cre+, Rac1f/f), heterozygous (Cre+,Rac1f/+), and control (Cre−,Rac1+/+) littermates. Upon collection, the lungs were either fixed and processed for embedding and histology, or were used for the extraction of RNA and gene expression analysis. Moreover, human lung explants were cultured in presence or absence of Rac1 inhibitor.

Results

Epithelial deletion of Rac1 resulted in lung hypoplasia starting at E13.5 and up to E18.5. During the early pseudoglandular stage the mutant lungs displayed dilated epithelial tips, and expressed high levels of Fgf10 and Wnt inhibitors. The mesenchymal deletion of Rac1 resulted in lungs that also appeared hypoplastic starting at E14.5 until E18.5. However, these mutant lungs displayed severe hypercellularity starting at E14.5. At E18.5, mesenchymal deletion of Rac1 resulted in severe reduction in saccular spaces. Transcription levels of Wnt inhibitors appeared decreased in the E14.5 mutants. Interestingly, both epithelial and mesenchymal Rac1 deletion resulted in embryonic lethality past E18.5. Using Wnt reporters, we showed that epithelial Rac1 deletion increased Wnt signaling, while mesenchymal Rac1 deletion decreased Wnt signaling. Moreover, we demonstrated that Rac1 is expressed in human fetal lung at different gestational stages. This expression is higher in the mesenchyme as compared to the epithelium. Treating human lung explants with Rac1 inhibitor resulted in impaired explant branching and growth.

Conclusion

In summary, both epithelial and mesenchymal Rac1 are required for proper lung development, with opposing effects on Wnt signaling. Moreover, Rac1 seems to play an important role in human lung development. The precise mechanism by which epithelial vs mesenchymal Rac1 act, and the differential signaling molecules it regulates in each compartment require further investigation.

Support or Funding Information

This work is supported by the AHA.
Effects of Corn Oil, Compared to Extra Virgin Olive Oil, Intake on Lipoprotein and Subfraction Cholesterol Levels in Men and Women: Results from a Randomized, Controlled Feeding Study
Authors: Maki Kevin C , Lawless Andrea L , Kelley Kathleen M , Kaden Valerie N , Geiger Constance J , Palacios Orsolya M , Dicklin Mary R
Published: November 2023

The aim of this analysis was to compare the effects of consuming foods prepared with corn oil (CO) versus extra virgin olive oil (EVOO) on fasting lipoprotein and subclass cholesterol levels in adults with elevated (130–199 mg/dL) low‐density lipoprotein cholesterol (LDL‐C) at baseline. A randomized, double blind, controlled‐feeding, crossover study design was used with an evaluable sample of 54 men and women with mean (SEM) baseline LDL‐C 147 (4.2) mg/dL. Subjects underwent two 21‐d controlled‐feeding periods, separated by a washout of ≥21 d. Feeding periods included 4 tbsp (~54 g) per day of CO or EVOO in 3 foods (muffin, dinner roll, yogurt). Fasting lipoprotein and subclass cholesterol levels were measured at baseline and at the end of each feeding condition. Density gradient ultracentrifugation (Vertical AutoProfile‐2 method, Atherotech Diagnostics Laboratory, Birmingham, AL) was used to assess total LDL‐C, small LDL3+4‐C, large LDL1+2‐C, intermediate‐density lipoprotein cholesterol (IDL‐C), total high‐density lipoprotein cholesterol (HDL‐C), large HDL2‐C, small HDL3‐C, total very low‐density lipoprotein cholesterol (VLDL‐C), large VLDL1+2‐C, and small VLDL3‐C. Total LDL‐C changed by a median of −8.6% during the CO condition versus −0.6% during EVOO feeding (P<0.0001). The larger reduction in total LDL‐C with CO was mainly attributable to changes (P<0.01 for CO vs. EVOO) in large LDL1+2‐C (−10.4%) and IDL‐C (−17.0%). CO feeding did not alter VLDL‐C, but EVOO raised the concentration by a mean of 10.7% (P<0.002 for CO vs. EVOO), due mainly to increased cholesterol in small VLDL3 particles (25.3%). Changes in both LDL‐C and VLDL‐C contributed to the more favorable median change in non‐HDL‐C with CO (−6.5%) compared with EVOO (0.8%; P<0.0001 for CO vs. EVOO). No significant differences between conditions in total HDL‐C, HDL2‐C or HDL3‐C responses were observed. In summary, foods made with CO produced more favorable effects than EVOO on atherogenic lipoprotein cholesterol (LDL, VLDL, non‐HDL) levels.

Support or Funding Information

Funded by ACH Food Companies, Oak Brook, IL and PepsiCo., Purchase, NY
Interplay between Substrate Stiffness and TGF‐b1 on Cellular Traction Force and Migration of MDA‐MB‐231 Cells
Authors: LIN PO‐YEN , Chu Yeh‐Shiu , Lai Yi‐Ting , Lo Chun‐Min
Published: November 2023

TGF‐β1 has been known to activate downstream signaling pathways of breast cancer cells and facilitate their invasive behaviors. In addition, our preliminary results show that MDA‐MB‐231 cells cultured on rigid substrates display higher traction forces and cell migration speed in comparison to cells cultured on more compliant substrates. The concept that a rise of contractile force due to increased ECM rigidity or TGF‐β1 induced scattering migration of MDA‐MB‐231 cells is intriguing, but the cross talk between ECM rigidity and TGF‐β1 remains to be elucidated. The goal of this study is to investigate the effect of substrate rigidity on TGF‐β1 induced changes of cell migration and traction forces of MDA‐MB‐231 cells. Collagen‐coated polyacrylamide substrates with a variety of Young's moduli (3.1, 7.4, and 10 kPa) were combined with fluorescence confocal imaging and digital image analysis to characterize a variety of cellular responses to substrate rigidity. Images of cell movement and the displacement of fluorescent beads embedded in polyacrylamide gels were taken every five minutes. Compared with control group without TGF‐β1 treatment, MDA‐MB‐231 cells on 7.4 kPa and 10 kPa gels demonstrated higher migration speed and smaller traction forces after TGF‐β1 treatment. However the addition of TGF‐β1 has little effect on traction forces or migration speed of MDA‐MB‐231 cells cultured on 3.1 kPa gels. Our results indicate that TGF‐β1 induced changes of scattering migration and traction forces are substrate rigidity dependent.

Support or Funding Information

This work was supported by the Ministry of Science and Technology, Taiwan.

(Grant number: NSC 102‐2628‐B‐010‐010‐MY3)
Fabrication of A Novel Elastomeric Substitute For Autologous Veins In Coronary Artery Bypass Surgery
Authors: Warner Harleigh J , Wagner William D
Published: November 2023

Autologous saphenous vein is the standard material for bypassing small diameter (<6mm) coronary arteries, but are subject to intimal hyperplasia, thrombosis, and accelerated atherosclerosis. To date, no biomaterial functions as a substitute for vein graft. We have recently developed an endovascular biomaterial, PFC, composed of electrospun poly‐(glycerol sebacate), silk fibroin, and type 1 collagen which has tensile, non‐thrombogenic, and endothelial cell adhesive properties ideal for an artery graft. In this study, PFC was tested to determine if enchanced mechanical properties and bioactive materials from cells would interface with PFC fibers to facilitate improved performance of the biomaterial. Sparse PFC mats composed of nanofibers were fabricated by electrospinning 200 μl of 10% PFC [(PGS: Fib: Col (1:4.5:4.5)] in 1,1,1,3,3,3‐hexafluoro‐2‐propanol (HFP) onto tissue culture plastic treated slides, resulting in a sparse network of PFC fibers. Samples were heat cured and crosslinked with gluteraldehyde. NIH 3T3 mouse fibroblasts were cultured on PFC fibers for 6 days and decellularized using Trypsin/Triton X‐100. Slides were imaged by SEM and light microscopy. Dense PFC mats were fabricated by electrospinning 4.4 mls of 10% PFC onto a 9 by 10cm rectangle of aluminum foil, heat curing, and crosslinking with gluteraldehyde. Fibroblasts were seeded on material for 10 days and either immediately fixed (n=3) or decellularized (n=3) and fixed. Materials were evaluated histologically. Decellularization protocols did not have an effect on the morphology of PFC fibers. Fibroblasts deposited a nanofibrous extracellular matrix (ECM) on the surface of sparsely electrospun PFC as shown by SEM and crystal violet staining. The ECM was apparent on sparce PFC after decellularization with Triton X‐100. The PFC facilitated the ability for the ECM to remain on the material compared to tissue culture plastic slides without PFC. Fibroblasts formed a dense monolayer on the surface of PFC. These studies show it is possible to effectively decellularize PFC seeded with fibroblasts and maintain ECM material, resulting in the ability to potentially tune the properties of the material.
Women's Empowerment and Food Security Status: A Global Comparative Study of Women Living in Rural and Urban Areas
Authors: Sinclair Kate , Ahmadigheidari Davod , Dallmann Diana , Melgar‐Quiñonez Hugo
Published: November 2023

In recent decades, many efforts have been made to empower women, nonetheless, many gaps in gender equality still exist. These gender disparities have negative consequences not only for women but also for their families and society as a whole. Empowering women plays a vital role achieving and maintaining a food secure state. The objective of this study was to examine the association between indicators that are considered to contribute to women's empowerment and food security (FS).

A sample of 15,334 rural women and 10,098 urban women was obtained from the 2015 Gallup World Poll, which is a nationally representative survey of individuals 15 years and older in 150 countries. The Food Insecurity Experience Scale within this dataset was used to categorize by FS status (i.e. Secure and Insecure) as the dependent variable, while women's empowerment indicators were used as independent variables. Crosstabs analyses and binary logistic regressions, for both rural and urban areas, were carried out using SPSS 23 Complex Sample.

Several socio‐demographic characteristics, including education, marital status, employment status and income, were found to show significant differences between women living in rural and urban areas by FS status. Also, rural women were at a higher risk of being food insecure. Findings from the logistic regression show that rural women with higher levels of social support (OR 0.697), education (OR 0.175), control over life (OR 0.513), and personal health index (OR 0.402), had significantly higher odds of being food secure. Similar results were found among women living in urban areas in terms of education (OR 0.241), personal health index (OR 0.209) and control over life (OR 0.445). Finally, urban women who perceived being treated with respect (OR 0.438) also had higher odds of being food secure.

The results confirm that the determinants of food security are highly complex, they do not act autonomously, and their effects are intertwined. Nonetheless, it is clear that factors contributing to women's empowerment play a key role in FS. Women's access to education is invaluable and should be prioritized worldwide as a means of improving FS. Shifting social norms to allow women to take control of life decisions is essential in achieving FS. Insuring one is able to maintain a good health status is pertinent to FS. As other studies have shown, strengthening social support is also a valuable tool to combat food insecurity, especially among rural women. Finally, living in urban areas provides increased and diverse opportunities for women compared to living rural areas. Addressing this opportunity gap in the future will be helpful in ensuring women's empowerment and FS for all.
Stromal Cell‐Derived Extracellular Matrix Stimulates Mesenchymal Stem Cell Proliferation and SSEA4 Expression When Used as Supplement in Cell Growth Medium
Authors: Zamilpa Rogelio , Flores Iris , Navarro Mary M , Griffey Sy
Published: November 2023

Mesenchymal stem cells (MSC) have been shown to promote tissue repair and regeneration due to their ability to immunomodulate the microenvironment, stimulate angiogenesis, and give rise to multiple cell types. However, one of the obstacles associated with MSC therapies is obtaining relevant numbers of MSCs for clinical treatments. In order to improve MSC isolation and expansion, we developed a High Performance Micro Environment (HPME) generated by bone marrow stromal cells. Scanning electron microscopy and mass spectrometry analyses demonstrate that the HPME is a three‐dimensional, protein network composed of extracellular matrix proteins. When used as a cell culture substratum, the HPME improves the yield of adipose‐, bone marrow‐, and cord blood‐derived MSCs by a minimum of 2‐fold. Bone marrow MSCs expanded on HPME have an increased expression of stage specific embryonic antigen (SSEA)‐4, a multipotent MSC marker, when compared to MSCs expanded on plastic (41±6% difference compared to plastic). Interestingly, an HPME protein extract used as a supplement in cell growth medium stimulated MSC proliferation and increased the MSC yield in a dose‐dependent manner. Concomitantly, MSCs stimulated with HPME extract also had an increased expression of SSEA‐4 (43±4% difference compared to unstimulated). These results indicate that the HPME is an ideal substratum or medium supplement for expansion of MSCs to be used in therapeutic applications.
The Critical Role of SENP1‐Mediated GATA2 DeSUMOylation in Graft Arteriosclerosis by Promoting Endothelial Activation
Authors: Qiu Cong , Wang Yuewen , Zhu Xiaolong , Song Lin , Zhang Haifeng , Qin Lingfeng , Tellides George , Min Wang , Yu Luyang
Published: November 2023

Immune rejection mediated graft arteriosclerosis (GA) is the major cause of graft failure after allograft transplantation. The pathological process of GA starts from endothelial cell (EC) activation marked with inducible expression of endothelial adhesion molecules, which results in neointima formation, vasomotor dysfunction and consequent vascular occlusion. However, the comprehensive regulatory mechanism of EC activation during GA remains unclear. In the current study, dramatic but endothelium‐specific induction of SENP1, the major protease of small ubiquitin‐like modifier (SUMO) system, was observed in clinical graft rejection specimens by immunofluorescence staining, synchronized with endothelial activation mediated vascular remodeling. Of note, GATA2, the master transcriptional factor regulating expression of adhesion molecules, had similar expression pattern as SENP1. According to the correlation of endothelial SENP1 in GA, SENP1 endothelial conditional knockout mice (SENP1‐ecKO) were generated and subjected to an aorta transplantation model of GA. Endothelial SENP1 deficient graft maintained a better vessel function and had limited leukocyte recruitment and neointima formation during GA progression. Induction of endothelial adhesion molecules in early stages of GA was clearly diminished in the absence of endothelial SENP1 and this effect was confirmed in cultured mice aortic ECs as well as human EC bearing SENP1 deSUMOylase inactive mutant, indicating the regulatory effect of SUMOylation on EC activation. Indeed, constitutive SUMOylation of endothelial GATA2 was identified in SENP1‐ecKO graft and cultured EC, which was directly mediated by inflammation induced SENP1. Mechanistically, SUMOylated GATA2 demonstrated weakened stability and DNA binding onto promoters of adhesion molecule genes, resulting in dampened EC activation and consequently leukocyte‐endothelial adhesion and transmigration. More importantly, endothelial overexpression of GATA2 reversed the retarded GA progression in SENP1‐ecKO graft, further supporting the critical axis of SENP1‐GATA2 in endothelial inflammation. Taken together, we conclude that inflammation mediated SENP1 boosts GA by regulating GATA2 SUMOylation and consequent EC dysfunction.

Support or Funding Information

This work was supported by National Natural Science Foundation of China (Grant No. 81270357, 81422005 and 31470057), and Zhejiang Provincial Natural Science Foundation of China (Grant No. LR14H020002)
Transcutaneous vagus and trigeminal nerve stimulation for neuropsychiatric disorders: underscoring anatomical landmarks and connections
Authors: Barros Mirna Duarte , Trevizol Alisson , Sato Isa , Liquidato Bianca Maria , Cordeiro Quirino , Shiozawa Pedro
Published: November 2023

We analyzed trigeminal nerve stimulation (TNS) and transcutaneous vagus nerve stimulation (tVNS). All techniques have shown preliminary promising results regarding different neuropsychiatric scenarios. Our group recently developed a new TNS protocol for different disorders such as depression, generalized anxiety, fibromyalgia, panic attacks and substance addiction. Considering all patients enrolled for our different studies, a total of 100 patients underwent the referred stimulation protocol despite clinical scenario. We used an external neurostimulator to deliver electric current transcutaneously through orbital branch of the trigeminal nerve. The parameters for this technique are frequency, pulse duration, train duration, density (intensity of stimuli/electrode area) and electrode position, revealing the importance of anatomical knowledge about trigeminal nerve branches, their position and connections. The protocol is as follows: frequency of 120 Hz, 200ms of pulse duration for 30 minutes. The intensity is determined individually and corresponds to the intensity that each subject refers as a non‐painful mild paresthesia without muscle contraction. The electrodes are 25cm2 and are positioned over supraorbital foramen with the negative (cathode) on the right side and positive (anode) on the left. A total of 10 sessions (one session a day) are performed. Regarding clinical outcomes, overall data analysis demonstrate a significant participation of TNS in ameliorating neuropsychiatric symptoms of patients enrolled for our trials and case studies. The technique was not related to any severe adverse effect. In fact, patients presented only a mild transient erythema following the active TNS stimulation, which would fade away in a few minutes. Skin injury is systematically assessed in every study protocol following the suggestion of Basketter and colleagues. The authors provide evidence that visual scoring can be sensitive reliable and reproducible within a study protocol. The present TNS protocol used for different clinical trials in neuropsychiatry is a safe strategy regarding skin injury, in contrast to similar non‐invasive neuromodulation strategy such as tDCS, which markedly present safety concerns regarding skin integrity in different reports. The present results, however significant, need to be taken as hypothesis‐driven. We understand that the trends seen so far shall strongly justify larger double‐blind studies in this area
Functionalizing the Unannotated Mitochondrial Proteome
Authors: Rutter Jared P.
Published: November 2023

Mitochondria are dynamic and complex organelles that play a central role in all aspects of biology, including energy production, intermediary metabolism, and apoptosis. These broad cellular functions also place mitochondria as a central player in human health. Mitochondrial dysfunction is associated with a wide range of diseases, including cancer, type 2 diabetes, and most neurodegenerative disorders. As a result of these wide‐ranging critical activities, many efforts have focused on identifying and characterizing the mitochondrial proteome, with over 1,000 proteins identified to date in mammals. Remarkably, however, roughly one‐quarter of these proteins remain essentially uncharacterized. These include many proteins that are highly conserved throughout eukarya, a strong indication that they perform a fundamentally important function. Our studies of a handful of these uncharacterized mitochondrial conserved proteins support this proposal, revealing new roles for these proteins in critical aspects of mitochondrial function, including mitochondrial protein quality control, lipid synthesis and mitochondrial ETC complex and supercomplex assembly. In two cases, these fundamental observations led directly to the elucidation of new human disease genes. One of these discoveries provided the first molecular identification of the protein complex that is necessary and sufficient for mitochondrial pyruvate entry. We have made progress in understanding the biochemical mechanisms governing the activity and structure of this complex. We have also found that loss of its activity is an important feature of cancer metabolism. The overall goal in this research is to provide a new understanding of the biochemical and cellular function of each conserved uncharacterized mitochondrial protein, determine how they contribute to normal mitochondrial activity and human disease.
The role of mammalian polymerase theta in DNA repair
Authors: Sfeir Agnel
Published: November 2023

Following telomere de‐protection chromosome ends are fused either by a Ligase 4‐dependent classical NHEJ process (C‐NHEJ) or using an alternative NHEJ pathway (alt‐NHEJ) mediated by Ligase 3 and PARP1. We recently investigated the differences in sequence fidelity between the two end‐joining reactions, and identified Polymerase theta (Polθ; encoded by Polq) as a critical alt‐NHEJ factor that introduces random nucleotides at fusion junctions. Polq inhibition suppresses alt‐NHEJ at telomeres that have lost shelterin protection, and blocks translocations at non‐telomeric loci. In parallel, loss of Polq results in increased rates of homology directed repair (HDR). Interestingly, depleting Polq in cells carrying mutations in the breast‐cancer susceptibility genes (Brca1 or Brca2) exacerbates chromosomal aberrancies and reduces cellular survival, suggesting that Polθ‐mediated alt‐NHEJ compensates when HDR is impaired.

Polθ is an atypical A‐family DNA polymerase with an N‐terminal helicase‐like domain, a large central domain harboring a Rad51 interaction motif, and a C‐terminal polymerase domain capable of extending DNA strands from mismatched or even unmatched termini. In vitro experiments identified an evolutionarily conserved loop in the polymerase domain that is essential for synapsing DNA ends during end joining. To address the mechanism by which this low‐fidelity polymerase orchestrates double‐strand break repair we employed CRISPR/Cas9 gene editing and established cell lines with key Polθ mutations (PolqΔhelicase, PolqΔRAD51, PolqΔpolymerase, and PolqΔloop). Here we report on the impact of these mutations on Polθ function in promoting alt‐NHEJ and suppressing HDR, and address the importance of Polθ‐mediated alt‐NHEJ in survival of HDR defective cells.
NMR Relaxation Dispersion
Authors: Loria Patrick
Published: November 2023

NMR relaxation dispersion experiments are used to characterize millisecond and microsecond motions and the role of these motions in regulation of protein tyrosine phosphatases (PTPs) In a pair of mechanistically indistinguishable PTPs with catalytic rates that vary by 50‐fold these experiments indicate that catalytic activity is regulated by the rate of active site loop closure. Furthermore, it is observed that small molecule binding distant from the active site loop alters the kinetic rates of substrate catalysis as well as the dynamics of loop closure.
Quality Control In 40S Ribosome Maturation
Authors: Karbstein Katrin
Published: November 2023

While much is known about how ribosomes faithfully translate mRNAs into protein, how ribosomes are assembled within cells remains poorly understood. With ~ 2,000 ribosomes synthesized every minute ribosome assembly is the major metabolic activity of a growing cell. Given its profound energy requirements, ribosome assembly is tightly regulated in response to extracellular stimuli. In addition to providing sufficient numbers of ribosomes, cells must also ensure that the produced ribosomes are correctly assembled. This is important to safeguard their catalytic activity and extraordinary fidelity. We are exploring cellular mechanisms to ensure proper assembly of nascent 40S subunits.

Support or Funding Information

R01 GM086451
MCAT2015: A New Focus On Biochemistry
Authors: Oyewole Saundra Herndon
Published: November 2023

The 5th comprehensive review of the Medical College Admissions Test (MCAT), informed by the AAMC‐Howard Hughes Medical Institute's “Scientific Foundations for Future Physicians (SFFFP)” and the AAMC “Behavioral and Social Science Foundations for Future Physicians” reports, resulted in major revisions to the MCAT. Both reports focused on the competencies that entering medical students need to be successful in medical school and in medical practice. Among the changes in the MCAT is the addition of biochemistry as one of the science content areas. Biochemistry concepts were added based on data presented in the SFFFP report, survey data from medical school respondents, and input from stakeholders at various outreach events regarding the importance of biochemistry. The data suggest that knowledge of biochemistry fundamentals is increasingly important to the academic success of entering medical students.

This presentation will provide an overview of the entire exam, with a focus on the importance of scientific inquiry and research skills as students apply their knowledge to answer questions based on passages describing research protocols. Students will be required to analyze data presented in various formats (tables, graphs, etc.) in order to answer passage‐based questions. This exam includes biochemistry content covered in many first semester biochemistry courses. The format of questions requires a deeper understanding of research methods and data analysis than the former MCAT. In addition, the new MCAT covers cellular and molecular biology content that is taught in introductory biology sequences. The speaker served on the MR5 Committee that made the recommendations for the new MCAT.
Adaptations To Mitochondrial Dysfunction Via Inter‐Organelle Communication
Authors: Haynes Cole M
Published: November 2023

During metazoan development and cell specification, mitochondrial biogenesis is tailored to match the physiologic requirements of individual cell types. Loss of mitochondrial activity, or mitochondrial dysfunction, is tightly linked with a number of seemingly disparate pathologic scenarios including neurodegeneration, cancer, diabetes and bacterial infection. My laboratory is interested in how cells evaluate mitochondrial function and adapt to survive and ultimately recover from mitochondrial stress. By studying a transcriptional response pathway known as the mitochondrial unfolded protein response (UPRmt), we identified ATFS‐1. Because this unique transcription factor has both a mitochondrial and nuclear localization sequence it can function as a sensor of mitochondrial function to couple defects in mitochondrial protein import efficiency and the activation of a broad mitochondrial protective transcriptional response. I will discuss protective roles for the UPRmt during a variety of physiologic conditions associated with mitochondrial dysfunction including during bacterial infection caused by pathogens that specifically perturb mitochondrial function to promote virulence.

Support or Funding Information

This work was supported by the Ellison Medical Foundation and NIH grants R01AG040061 and R01AG047182.
Creating Stop‐Motion Animations To Learn Molecular Biology Dynamics
Authors: Peterson Celeste , Ngo Pauline
Published: November 2023

Classes in molecular biology have historically used static diagrams to depict molecular processes where arrows represent changes. However, these cartoons do not capture how molecular processes occur in time and 3D space, and the large number of arrows and steps is often confusing. Here we describe 3D stop motion animation as an alternative. Students choose a molecular process, plan how to represent it with physical objects, and produce a stop motion video that is shared with other students over the internet. These animations can be easily implemented in courses and require very little resources. We have found that students enjoyed making them and especially sharing them with their peers. They are appropriate for undergraduate biology courses at all levels ranging from introductory courses to upper level research labs where students create animations of their own findings. Overall, this exercise offers the students a multisensory learning experience and the opportunity to contribute to open education resources.