bluuebunny/crossref_metadata_2025 huggingface dataset:
The effect of shifting food‐intake period on rat body weight and plasma levels of glucose, leptin, melatonin, cholesterol and triglyceride
Authors: Tanira Musbah Omar , Al Qutiti Abdulmajeed , Al Jahwari Basim
Published: June 2021

The suprachiasmatic nucleus of the hypothalamus receives photic information from the retina and contribute to the control of body functions and hormonal secretion in a rhythmic manner through day/night cycle. We hypothesized that disturbance of circadian rhythm by shifting food‐intake period may reflect on body metabolism‐related blood parameters and hence body weight. Hence, we investigated the effect of repeated shifting food intake (over a period of six weeks) on daily changes of rat body weight, food/water intake, feces output and plasma levels of glucose, leptin, melatonin, cholesterol and triglyceride. Two groups of male WKY rats weighing 150–180g were used. The first was subjected to a constant day/night of 12:12h of light/dark cycle (fixed light/day cycle throughout the experimental period; Fixed Group); whereas the second group was subjected to an alternating (every two weeks) pattern of light/dark‐dark/light cycle of 12:12h each (Shifted Group). The plasma glucose, triglycerides, leptin and melatonin levels as well as feces output, food intake showed significant differences between Shifted and Fixed Groups. However, body weight and other measured plasma parameters were not significantly different. It is concluded that rats may need more frequent shifting of food‐intake period to manifest more conclusive effects.
Wound healing effect of TENS in rodents.
Authors: Barreto André Sales , de Souza Siqueira Barreto Rosana , de Souza Siqueira Quintans Jullyana , de Oliveira Donato Isabela , Cardoso Roberta Santana Deodorio , Soares Rosilene Calazans , Albuquerque‐Júnior Ricardo Luiz Cavalcante , de Lucca Waldeci , de Santana Josimari Melo , Quintans‐Júnior Lucindo José
Published: June 2021

The aim of this study was to investigate the wound healing activity of low frequency and high frequency TENS on excision wound model in rats.

Excision wound:

Thirty rats, Wistar, 250–300 g were anesthetized and an excisional wound was performed. The animals was treated with placebo ‐ TENS simulation (equipment off), low‐frequency TENS (2 Hz) and high‐frequency TENS (100 Hz) for 7 days. The residual wounds were analyzed and removed. Experimental protocols were approved by the animal care and use committee at Federal University of Sergipe (CEPA 69/2011). Serial histological sections were obtained and stained by histochemical techniques (Hematoxilin‐Eosin – HE and Sirius red) and immunohistochemical techniques. The results suggest that the high‐frequency TENS reduced (p < 0.05) wound areas. This evidence shows that high‐frequency TENS modulates the wound healing process.
The Role of α‐and β‐Estrogen Receptors on Neurite Outgrowth in Neuro‐2A cells
Authors: Deshotels Matthew Risher , Spinu Eduard , Bailey Evangeline , Filipeanu Catalin
Published: June 2021

Although the neuroprotective actions of estrogens have been well established, the mechanisms underlying these effects remain obscure. Thus, we aimed to identify the estrogen receptor (ER) subtype involved in neurite outgrowth, a marker of neuroprotection. To this end, myc‐tagged α‐and β‐ER were transiently transfected in Neuro‐2A cells which lack endogenous expression of both receptor subtypes, as demonstrated by western blot. The expression of both α‐ER and β‐ER resulted in significant increases in the number of cells bearing neurites (25 ± 6.1% and 31± 8%, respectively) compared to control cells (15 ± 3.3%). However, treatment with the ER agonist 17β‐estradiol (10 nM) did not further enhance the number of neurites (29 ± 6.3% for α‐ER and 28 ±7.2% for β‐ER). The average length of extended neurites was also increased by expression of α‐ER and β‐ER (59 ± 5 μm and 63 ± 7 μm, respectively) compared to controls (49 ± 6 μm). The neurite outgrowth observations were correlated with cellular BDNF levels. In conclusion, at least in the present heterologous transfection system, constitutive signaling activity of both ER subtypes accounts for neuronal differentiation.
Prenatal Cocaine Exposure Inhibits Neurite Outgrowth Through RAGE Pathway and Activates Altered Expression of Pro‐addictive Cytokines in Primary Neuronal and Glial Cell Cultures
Authors: Johnson Marquita L , Womble Tracy , Jackson Tanise L
Published: June 2021

Background

The prevalence of neonates exposed to cociane continues to be an endemic within the U.S. An overwhelming amount of evidence has shown that prenatal cocaine exposure is associated with numerous neurological and behavioral abnormalities, as well as structural damage and a deficiency in neurodevelopment with no established underlying mechanisms.

Purpose

The present study was designed to investigate biochemical events coupled to cocaine‐induced cytotoxicity of neurons and glial cells seen in exposed offspring.

Research Approach:

Time pregnant Sprague‐Dawley rats were treated with 40 mg/kg of cocaine HCl on gestational day (GD) 10–21. On postnatal day (PND) 3 the brain tissue of the pups was harvested to yield mixed neuronal and glial cell cultures. A cytokine antibody array determined a decrease in RAGE expression and an increase in TIMP‐1 expression in prenatally exposed pups. Immunocytochemistry (ICC) showed decreased neurite outgrowth in prenatally exposed neurons. ICC also determined there was an increase in dopamine (DA) in prenatally exposed primary cell cultures indicating the presence of DA receptors on astrocytes and oligodendrocytes.. Western Blot analysis quantitatively determined a variation of amphoterin (HMGB1) which is necessary for RAGE activation and pro‐inflammatory cytokine matrix metalloproteinase 9 (MMP‐9) protein expressions between the control and cocaine treated pups.

Conclusion

Through in vivo and in vitro studies exposure to cocaine during gestation leads to marked alterations in morphology and function of primary neuronal and glial cell cultures. These findings suggest that exposure to cocaine in utero can ultimately increase the incidence of addiction and development of neurodegenerative diseases and neuropathologies in the affected offspring.
Palmitoylethanolamide Enhances Brain‐Derived Neurotrophic Factor Production and Neurogenesis in the Hippocampus Following Ischemic Brain Injury
Authors: Cuzzocrea Salvatore , Crupi Rosalia , Paterniti Irene , Impellizzeri Daniela , Campolo Michela , Esposito Emanuela
Published: June 2021

Stroke is the third leading cause of death and the leading cause of long‐term disability in adults. To counteract the ischemic brain injury, a new therapeutic approach has been employed by using palmitoylethanolamide (PEA). Ischaemic injury was induced for 2 h by middle cerebral artery occlusion (MCAo) followed by 24‐h reperfusion. PEA (10 mg/kg, i.p.) was administered by intraperitoneally injection twice, at 1 after the onset of ischemia and 6 h after reperfusion. In the present study using a transient ischemia model, we showed that (1) PEA induced the phosphorylation of extracellular signal‐regulated kinases 1/2 (ERK1/2) and cAMP response element‐binding protein (CREB) in the hippocampus after ischemia; (2) PEA increased the expression of brain‐derived neurotrophic factor (BDNF), a representative neurotrophic factor in the central nervous system, in the hippocampal dentate gyrus, and most BDNF‐positive cells were also stained with anti‐glial fibrillary acidic protein (one of the major intermediate filament proteins of mature astrocytes) and (3) PEA increased doublecortin positive neuronal precursor cells in the dentate gyrus subventricular zone or subgranular zone. These results suggest that PEA has the ability to induce BDNF production in astrocytes and enhance neurogenesis after brain ischemia, which may be mediated by activation of ERK1/2 and CREB.
Bioinformatic analysis of microglia‐neural stem cell interactions: a role for wnt5a?
Authors: Woodbury Maya , Ingraham Kaitlin , Freilich Robert , Ikezu Tsuneya
Published: June 2021

In the adult mammalian brain, neural stem and progenitor cells (NSCs, NPCs) persist in the subventricular zone (SVZ) and the hippocampal subgranular zone (SGZ). The SGZ microenvironment contains various cell types in close association with NSCs, and signaling within this environment regulates maintenance, proliferation and differentiation of NSCs. Signaling from astrocytes to neurons in hippocampal neurogenesis has been described, but factors neurons receive from microglia are rather unexplored.

Using genome‐wide microarray analysis, we assessed gene expression in microglia and embryonic NSCs co‐cultured with conditioned media from microglia (MCM) treated to induce pro‐inflammatory (M1) or protective (M2) conditions.

We identified Wnt5a as a growth/differentiation factor highly expressed by microglia. In NSCs, we observed differential expression of Wnt5a target genes in each condition. NSCs co‐cultured with M2 MCM showed upregulation of growth/differentiation‐related genes vs. those with M1 MCM. Wnt5a knockdown in microglia was associated with decreased NR2B expression in co‐cultured NPCs vs. control, implying that Wnt5a is an essential microglia‐derived factor for synaptic maturation. These data suggest that microglia play a key role in regulating neurogenesis, and highlight the Wnt5a microglia‐neuron pathway as a potential target for neurodegeneration and developmental disorders.
Role of Voltage‐dependent Kv7 Channels in the Regulation of Coronary Blood Flow
Authors: Goodwill Adam G , Casalini Eli D , Owen Meredith K , Conteh Abass , Sassoon Daniel , Shatagopam Kashyap , Dick Gregory M , Tune Johnathan D
Published: June 2021

Previous studies have demonstrated a key role for voltage‐dependent K (KV) channels in the regulation of coronary metabolic and ischemic vasodilation. However, the KV channel subtypes responsible for the control of coronary blood flow have not been clearly defined. This investigation examined the hypothesis that KV7 channels are present in coronary vascular smooth muscle and regulate perfusion of the myocardium at rest and during pacing‐induced increases in metabolism. In anesthetized, open‐chest swine, we found that intracoronary administration of the KV7 agonist flupirtine (1 nM‐10 μM) or the KV7 antagonist linopiridine (1–100 μM) did not significantly affect baseline coronary blood flow (Δ = −0.002 ± 0.005 vs. −0.010 ± 0.004 ml/min/g, respectively). H2O2 increased coronary flow ~2‐fold and this response was unaffected by linopiridine. Whole‐cell recording of isolated coronary smooth muscle cells revealed that flupirtine and linopirdine both inhibited KV current by ~25 ± 3% (P < 0.05). Cardiac pacing (57 + 5 to 102 + 12 bpm) increased myocardial oxygen consumption (~35%) and coronary blood flow (~38%). However, linopridine (10 μM) did not significantly alter this increase in coronary flow or the slope of the relationship between coronary blood flow and myocardial oxygen consumption (P = 0.61). These data do not support a prominent role for KV7 channels in the regulation of coronary blood flow. (Support: HL092245)
Over‐expression of FXRG and miR‐1 increases formation of RISC complexes in H9C2 cell line
Authors: Walker Lisa , Dockstader Karen , Slavov Dobromir , Sucharov Carmen
Published: June 2021

miRNAs regulate gene expression by binding to mRNAs and triggering the formation of the RISC complex, which can promote mRNA degradation or translation repression in association with RNA binding proteins. Previous studies have shown that the Fragile X Related Protein 1 (FXR1) can modulate miRNA function. Undifferentiated H9C2 cells do not express the cardiac isoform of FXR1, whereas these isoforms (G and E) are expressed in differentiated cells. In undifferentiated cells miR‐1‐mediasted repression of connexin 43 (Cx43) is dependent on expression of FXR1G or E. To determine if FXR1G was involved in the loading of Cx43 in the Risc complex, H9C2 cells were co‐transfected with a DNA construct containing the Cx43 3′UTR (contains the miR‐1 binding site) linked to 24 MS2 binding sites and a YFP‐MS2 binding protein construct. The cells were infected with a construct over‐expressing FXRG‐FLAG and miR‐1. To quantify the mRNA bound to the RISC complex, immunoprecipitation was performed with AGO2 (a component of the RISC complex), YFP and FLAG antibodies. RNA was extracted and analyzed by qRT‐PCR. In undifferentiated H9C2 cells, FXRG and miR‐1 over‐expression results in increased binding of Ago2 to the Cx43 3′UTR. These results suggest that FXR1G is necessary for loading of the target mRNA to the RISC complex. Supported by the American Physiological Society Undergraduate Summer Research Fellowship.
STRUCTURAL REMODELLING IN THE LEFT VENTRICLE OF STREPTOZOTOCIN ‐INDUCED TYPE 1 DIABETIC RAT
Authors: Singh Jaipaul , Bidasee Keshore , Moore Caronda , Iqbal Tehreem , D'souza Alicia
Published: June 2021

Heart failure in diabetes mellitus is a major cause of premature morbidity and mortality but the underlying mechanisms are elusive and treatment remains empirical. This study investigated the role of chronic hyperglycemia (HG) in the development of hypertrophy and fibrosis in the left ventricle (LV) of streptozotocin (STZ)‐induced type 1 diabetic (T1DM) rats compared to age‐matched Wistar controls. T1DM rats and their hearts weighed significantly (p<0.05) less than age‐matched controls along with a reduction in the heart weight/body weight ratio. Plasma glucose levels were 5–6 folds higher in T1DM animals. Visual analysis of the stained LV sections revealed general disorganization of myocardial architecture characterized by thinner fibers, disarray of myofibers and scarcity of myofibrils in the T1DM group compared to control. The width and diameter of myocytes from the T1DM group were significantly less compared to age‐matched controls. In addition, the LV of T1DM rats were infiltrated with increased ECM deposition in both interstitial and perivascular regions and area of interstitial fibrosis was significantly greater in T1DM hearts compared to controls. LV of T1DM rats exhibited signs of end stage apoptosis, manifesting as condensation of nuclei and cell shrinkage. The results show that chronic HG can produce marked alterations in myocardial architecture characterized by fibrosis proliferation, apoptotic cell death and atrophy of myocytes. Supported in part by University of Central Lancanshire and NIH HL085061.
Endothelial nitric oxide synthase S‐glutathionylation and Ser1177 phosphorylation modulate myocardial protection
Authors: Talukder M A Hassan , Yang Changjun , Varadharaj Saradhadevi , Chen Chun‐An , Zweier Jay L.
Published: June 2021

Endothelial nitric oxide synthase (eNOS) plays an important role in cardiovascular function, myocardial injury and protection. The role of eNOS is well recognized in ischemic preconditioning (IPC); however, questions remain regarding how eNOS is regulated during ischemia‐reperfusion (I/R) and IPC. eNOS phosphorylation at Ser1177 enhances enzyme activation and nitric oxide (NO) production; however, S‐glutathionylation of eNOS uncouples the enzyme leading to loss of NO production and vascular dysfunction. We measure myocardial eNOS S‐glutathionylation and phosphorylation in the process of I/R injury and investigate the role of these modifications in cardioprotection. Isolated rat hearts were subjected to 30‐min of global ischemia and/or reperfusion with or without IPC stimuli. Ischemia induced a 4–5 fold increase in eNOS S‐glutathionylation that was completely reversed by â‐mercaptoethanol. Importantly, IPC reduced I/R‐induced eNOS S‐glutathionylation by 50%, decreased myocardial superoxide generation by >;80% and attenuated myocardial infarction by 75%. Levels of eNOS phosphorylation at Ser1177 decreased with I/R but were increased by IPC. Thus, IPC stimuli inhibit I/R‐induced eNOS S‐glutathionylation but increase Ser1177 phosphorylation to maintain eNOS coupling, activation and NO production that protect myocardium against I/R injury. (Support: NIH grants HL63744, HL65608 and HL38324).
Acute oral amino acid intake increases both secretion and breakdown of very low density lipoprotein‐triacylglycerol
Authors: Hurren Nicholas Michael , Porter Craig , Chondronikola Maria , Yfanti Christina , Volpi Elena , Belalcazar Ligia Maria , Kinsky Michael , Sidossis Labros , Børsheim Elisabet
Published: June 2021

We have previously reported a reduced plasma triacylglycerol (TAG) concentration in elderly individuals following chronic intake of amino acids (AAs), but it is unclear how acute intake of AAs affects plasma TAG kinetics. We used dual stable isotope tracer methodology in this study to investigate the effect of acute AA intake on the fractional secretion rate (FSR) and fractional breakdown rate (FBR) of VLDL‐TAG derived from plasma fatty acids. Eight hypertriacylglycerolaemic volunteers (mean ± standard deviation (SD): age 63.4 ± 7.7 years; BMI 29.6 ± 5.0 kg·m−2; fasting TAG 191 ± 52 mg·dl−1) were studied during a 5‐h basal period and a 5‐h period of acute AA intake. Infusions of U‐13C16‐palmitate and 2H2‐palmitate were started and stopped at specific time points to allow VLDL‐TAG secretion and breakdown to be calculated. VLDL‐TAG FSR was significantly higher during the AA intake period than during the basal period (mean ± SEM: 12.9 ± 2.6 vs. 8.7 ± 1.4 %·h−1; P < 0.05), as was VLDL‐TAG FBR (13.8 ± 4.0 vs. 4.9 ± 1.6 %·h−1; P < 0.05). We conclude that acute amino acid intake increases fractional secretion and breakdown rates of VLDL‐TAG, and may promote a state of net breakdown, which lowers the plasma TAG concentration. The study is sponsored by 1R01 AG033761 NIH/NIA and P30 AG024832 NIH/NIA. Screenings and study visits are being undertaken within the ITS‐Clinical Research Center at UTMB, funded by grant 1 U54RR026141 NIH/NCRR.
Neutralization of TNFα improves endothelial function and reduces vascular calcification in advanced atherosclerosis
Authors: Roos Carolyn M , Hagler Michael A , Kafa Rami , Zhang Bin , Miller Jordan D
Published: June 2021

Endothelial dysfunction and vascular calcification are major problems associated with prolonged hypercholesterolemia and aging, and are strongly associated with increases in TNFα. We hypothesized that inhibition of TNF‐α would reduce inflammatory and pro‐osteogenic gene expression in aorta, leading to improved endothelial function and reductions in intimal plaque calcification. We used hypercholesteremic mice (Ldlr−/−, apoB100/100) fed Western Diet for 6 months, at which point they received either intraperitoneal injections of saline (CTRL) or Infliximab (INX) for 3 additional months. Expression of the pro‐inflammatory genes TNF‐α, VCAM, and ICAM were all reduced in the INX group (qRT‐PCR). INX treatment improved endothelium‐dependent relaxation compared to the CTRL group (acetylcholine responses in isolated organ chamber; 50.7±3.4%, 43.7±3.7%; respectively), but treatment did not affect relaxation to nitroprusside. Furthermore, expression of the osteogenic gene MSX2 was reduced in INX mice, PPARγ target gene expression was increased, and intimal plaque calcification was reduced in aorta (Alizarin Red; CTRL = 13.7±2.9%, INX = 5.4±1.3%, respectively). In conclusion, blockade of TNF‐α may be a viable therapeutic target to reduce inflammation, improve endothelial dysfunction, and attenuate vascular calcification in the advanced atherosclerotic lesions commonly found in aging humans.

NHLBI
Lung fluid regulation at high‐altitude
Authors: Taylor Bryan , Summerfield Doug , Issa Amine , Kasak Alex , Johnson Bruce
Published: June 2021

Background

Whether lung fluid accumulates in acclimating individuals at high altitude (HA) remains controversial.

Purpose

To assess the effect of HA on lung fluid regulation.

Methods/Results

Systolic pulmonary artery pressure (sPAP) (echo), lung diffusing capacity for CO (DLCO), pulmonary capillary blood volume (Vc), alveolar‐capillary conductance (Dm) (single breath method), ultrasound lung comets (ULCs) and pulmonary function (PFTs) were assessed in 8 healthy adults at sea‐level and after 1 and 9 days at Mt. Everest Base Camp (EBC), Nepal (5150 m). Also, after 5 days at EBC, DLCO and PFTs were assessed in 5 subjects pre‐and post‐acute albuterol administration. EBC was reached after an 8 d hike at progressively increasing altitudes starting at 2800 m. Altitude exposure increased sPAP, had no effect on DLCO, increased Dm and decreased Vc with a subsequent increase in Dm/Vc, and decreased the number of ULCs (Table 1). There was no change in FVC or FEV1 but an increase in MEF50. Albuterol administration increased PEF (11.85±1.52 vs. 12.40±1.45 L/s) and MEF50 (4.85±0.77 vs. 6.00±1.30 L/s) but had no effect on DLCO, Dm or Dm/Vc.

Conclusion

These data suggest that lung fluid is decreased in acclimating healthy humans at high altitude.

Funding:

Mayo Clinic & The North Face.
Mitochondrial Dynamics in the Type 2 Diabetic Heart
Authors: Gonzalez Eileen R , Brito Alfonso , Peltekian Lila , Lopez Vanessa , Madala Sridhar , De La Fuente Christine , de Leon Ray , Yamazaki Katrina
Published: June 2021

Type‐2 diabetes (T2D) leads to an imbalance of mitochondrial fusion/fission events. Epicatechin is a cocoa plant flavanol that has shown cardioprotective effects in the ischemia reperfusion setting. Our objectives are to observe the protective effects of epicatechin in and in vitro and in vivo models of type 2 diabetes. In vitro, C2C12 myoblasts were cultured in glucolipotoxicity treatment (GLT) for 3 days in high glucose (~33 mM) and high fatty acids (0.6 mM) media, ± epicatechin. Our in vivo rat model included controls, sham ± epicatechin (1 mg/kg/day), and diabetes ± epicatechin (1 mg/kg/day). T2D was induced in vivo by high energy diet (HED) of 20% lard and 10% glucose. Low dose streptozotocin (30 mg/kg) injection was given at four weeks to decrease insulin secretion, after which epicatechin treatment began with maintained HED. In vitro results thus far show GLT average cell counts of 7.54E+04, which show significant increase in cell death compared to control (3.56E+05). Cells maintained in GLT setting with 100 nM epicatechin yielded an average cell count of 1.78E+0, thus demonstrating a reduction in cell death. Western analysis show enhanced mitochondrial proteins with epicatechin treatment compared to diabetic controls. We predict further data analysis will show a positive correlation between epicatechin and the restoration of normal mitochondrial dynamics in the T2D setting.
Mechanisms of population level variation in fatness and leanness
Authors: Wells Ann , Das Suchita , Ji Bo , Voy Brynn
Published: June 2021

Obesity represents the extreme of fat deposition. Adipose tissue from many obese models has been well‐characterized. However, much less is known about differences in adipose tissue between leaner and fattier individuals within a population. The objective of this study was to determine if fatness can be attributed to changes in expression of key lipogenic and adipogenic genes, in the context of a heterogeneous genetic background. The C57BL/6J * DBA/2J (BXD) recombinant inbred mouse strain panel was used to select 2 mice each from 6 lean and 6 fatty strains, based on percent abdominal fat. Expression of adipogenic and lipogenic genes were profiled using PCR Arrays (sabiosciences.com). Using Pearson's correlation genes correlated with adiposity were identified, and Student's t‐test was used to identify differentially expressed genes between lean and fatty mice. Ten of the 84 genes measured differed significantly between groups (p<0.05) including expected genes such as leptin and adipsin as well. Other genes less characterized with regard to fatness also differed between groups, including Cdkn1a and Mapk14 (p<0.05). Significant correlations were identified among adipokines and adipogenic genes independent of fatness, suggesting concerted regulation of adipogenic and endocrine functions. These data highlight pathways that alter adipose tissue composition and function in the absence of overt obesity.
Suppressing Allergic Airway Disease with Nitrated Fatty Acids
Authors: Reddy Aravind T , Lakshmi Sowmya P , Dornadula Sireesh , Pinni Sudheer , Rampa Dileep R , Reddy Raju C
Published: June 2021

Nitrated fatty acids (NFAs) are endogenous agonists of peroxisome proliferator‐activated receptor γ (PPAR‐γ) that have been shown to exhibit anti‐inflammatory and antioxidant activity. We investigated NFA effects in the OVA model of allergic airway disease. The glucocorticoid fluticasone (Flut) was used as a positive control. Pulmonary administration of the NFA 10‐nitro‐oleic acid (OA‐NO2) or Flut produced significant and essentially indistinguishable reductions in airway hyperresponsiveness, lung and BAL fluid cytokine concentrations, OVA‐specific IgE concentrations, and mucus production. Both likewise produced similar and significant reductions in eosinophil infiltration and oxidative stress, reflecting reduced adhesion molecule expression. In vitro studies using both immortalized and human bronchial epithelial cells cultured at an air‐liquid interface demonstrated that OA‐NO2 increased expression and activity of PPAR‐γ and decreased activity of the pro‐inflammatory transcription factor NF‐κB. Increased expression of the glucocorticoid receptor and activity of the antioxidant transcription factor Nrf2 were also observed. Our results thus demonstrate beneficial NFA effects that suggest potential therapeutic utility in asthma. This work was supported by National Institutes of Health grant HL093196.
Regional Wnt signatures in the colon and the influence of commensal bacteria
Authors: Neumann Philipp Alexander , Koch Stefan , Hilgarth Roland , Perez‐Chanona Ernesto , Jobin Christian , Parkos Charles A , Nusrat Asma
Published: June 2021

Wnt signaling is a key regulator of intestinal epithelial homeostasis. Since the large intestine exhibits considerable morphological and functional heterogeneity along its length, we investigated whether this can be explained by local differences in Wnt signaling. Furthermore, we examined whether divergent Wnt signaling may be shaped by commensal bacteria. Using a Wnt PCR array, expression of 84 genes related to Wnt‐mediated signal transduction was analyzed. A differential signature of Wnt related genes in the proximal vs. distal murine colon was identified. Specifically, we observed that Wnt agonists Wnt5a, Wnt8b, and Wnt11, Wnt receptors Fzd3 and Lrp6, and a Wnt inhibitory factor Wif1 were differentially expressed along the colon length. These Wnt signatures were associated with differential epithelial cell proliferation and migration in the proximal vs. distal colon. Given the importance of luminal bacteria in controlling epithelial homeostasis, Wnt gene expression in colonic segments of mice raised in germ‐free conditions were analyzed. Altered expression of Wnt genes was identified in mice raised in germ‐free vs. specific pathogen free conditions. These results suggest that the morphological and physiological heterogeneity within the colon is in part facilitated by differential expression of Wnt signaling components and influenced by colonization with bacteria.
Effect of dietary protein on bone status in US Adults aged 50 years and older; NHANES 1999–2004
Authors: Kim Jung Eun , Zanovec Michael , Fulgoni Victor L , Campbell Wayne W
Published: June 2021

Dietary protein is an important determinant of bone health, but the effect of dietary total protein intake (Tpro) on bone status of older adults is controversial. We assessed the effect of TPro on bone status using data from the 1999–2004 NHANES. Dietary (24‐h recall) and bone (DXA) data from 6,149 men and women aged 50+ y were analyzed. Subjects were grouped based on TPro (LP (n=1993), <0.8; MP (n=1150), 0.8–1.0; and HP (n=3006), >;1.0 g protein·kg ideal body weight (IBW)−1·d−1). Total‐body bone mineral content (BMC), appendicular BMC (aBMC), total‐body bone mineral density (BMD), and total‐body bone mass index (BoMI) were used as indicators of bone status. Mean protein intakes of LP, MP, and HP were: 0.58±0.004, 0.90±0.002, and 1.45±0.011 g protein·kg IBW−1·d−1, respectively. Compared to the LP, MP and HP had higher BMC (p<0.001), aBMC (p<0.001), and BMD (p<0.001) and HP had higher BoMI (p<0.001). There were no differences between MP and HP for these parameters. After covariate adjustment, HP had higher BMD (1.096 ± 0.003 g/cm2 vs. 1.085 ± 0.004 g/cm2; p < 0.05) than LP, with MP intermediate (1.094 ± 0.005 g/cm2). These results indicate inadequate protein intake adversely affects bone status in older adults. However, among subjects who consumed adequate protein, i.e. greater than 0.8 g protein·kg IBW−1·d−1, bone status was not influenced by higher protein intake.

Funding: Dairy Res Inst, Purdue Ingest Behav Res Ctr
Mesenchymal stem cells (MSC) from porcine bone marrow (pBM) of the Os femoris and the Crista iliaca
Authors: Stock Peggy , Brückner Sandra , Tautenhahn Hans‐Michael , Pelz Sandra , Hempel Madlen , Christ Bruno
Published: June 2021

MSC increasingly gain attention for clinical cell therapy of liver diseases. Before translation into the clinics feasibility must be proven preferably in large animal models.

The aim of the study was to demonstrate in the large animal model of the pig whether MSC from bone marrow (pBM‐MSC) of the Crista iliaca and from BM of the femur/shank display differences in regard to their hepatogenic differentiation potential.

MSC were isolated either from BM of the Crista iliaca or Os femoris. Mesenchymal features were analysed by flow cytometry. Functional properties like the synthesis of urea and glycogen were compared during hepatogenic differentiation.

pBM‐MSC from both sources were negative for hematopoietic markers CD14, CD45 but positive for mesenchymal markers CD44, CD29, CD90, CD105. pBM‐MSC from both sources could be differentiated along the adipocyte, osteocyte and hepatocyte lineage. With differentiation progressing glycogen and urea synthesis increased featuring typical hepatocyte functions. After 21 days of differentiation urea synthesis amounted to 3 nmol/10.000 cells × 24h comparable to the rate in primary pig hepatocytes after 8 days of culture.

pBM‐MSC from various sources are not different in respect to their mesenchymal features and their hepatogenic differentiation potential. Hence, hollow bones might be particularly useful to isolate allogeneic bone marrow cell transplants.
Excess postexercise leucine ingestion enhances muscle protein synthesis in skeletal muscle of older men
Authors: Dickinson Jared M , Gundermann David M , Walker Dillon K , Reidy Paul T , Borack Michael , Drummond Micah J , Arora Mohit , Volpi Elena , Rasmussen Blake B
Published: June 2021

Essential amino acid (EAA) ingestion enhances postexercise muscle protein synthesis, and in particular leucine is thought to promote a greater stimulatory effect. The purpose of this preliminary study was to determine the influence of ingesting excess leucine shortly following resistance exercise (RE) on skeletal muscle mTORC1 signaling and protein synthesis in older adults over a 24h time‐course. During a stable isotope infusion trial, older men performed a bout of RE and at 1h postexercise ingested 10g of EAA containing either a leucine content similar to high quality protein (CTRL, 1.8g leucine, n=6) or excess leucine (LEU, 3.5g leucine, n=6). Muscle biopsies (vastus lateralis) were obtained at rest and at 2, 5, and 24h postexercise to examine mTORC1 signaling and muscle protein synthesis. mTORC1 signaling was increased above rest in both groups at 2 and 5h, however, this increase was more pronounced in LEU at 5h. mTORC1 signaling returned to resting levels at 24h in both groups. Muscle protein synthesis was increased above rest in both groups at 5h, and remained elevated above rest at 24h in both groups. However, the increase in muscle protein synthesis at 5h was more pronounced in LEU. These preliminary data suggest that excess leucine ingestion following RE may enhance the acute anabolic response to resistance exercise in older adults.

NIAMS R01AR049877, NIA P30AG024832, NIH T32HD07539, NCATS UL1TR000071
Linoleic acid suppresses cholesterol efflux and ATP‐binding cassette transporters in murine bone marrow‐derived macrophages
Authors: Spartano Nicole L , Lamon‐Fava Stefania , Matthan Nirupa R , Obin Martin S , Greenberg Andrew S , Lichtenstein Alice H
Published: June 2021

Background

Individuals with diabetes are at increased cardiovascular disease (CVD) risk, possibly associated with elevated plasma free fatty acids (FA). Evidence suggests that elevated unsaturated (U)FA suppress macrophage cholesterol efflux, a mechanism implicated in CVD risk. Murine bone marrow‐derived macrophages (BMDM) were used to explore the relationship between elevated UFA and cholesterol efflux mediated by ATP‐binding cassette transporters (ABCA1 and ABCG1) and transcription factors liver‐x‐receptor‐alpha (LXR‐α) and sterol receptor element binding protein (SREBP)‐1.

Methods/Results

BMDM were stimulated with 100 uM linoleic acid (LA) or palmitic acid (PA) for 16 hrs, and exposed to 25 ug/mL oxidized low density lipoprotein (oxLDL) for 24 hr. LA and PA each suppressed ABCA1 mRNA (60% and 30%, respectively) and ABCG1 mRNA (54% and 29%, respectively) (all p<0.01). LA decreased high density lipoprotein (HDL) mediated cholesterol efflux by 53% and ABCA1 protein by 94% (both p<0.05) with no significant effect on ABCG1, LXR‐α or SREBP‐1 protein.

Conclusion

These results suggest that elevated LA attenuates macrophage HDL‐mediated cholesterol efflux through alterations in ABCA1 mRNA and protein but not by regulating LXR‐α or SREBP‐1 expression. This mechanism may contribute to diabetes‐associated CVD.

NHLIBI‐T32‐HL069772, USDA 58–1950‐0–0014
Linking obesity with colorectal polyp risk
Authors: Comstock Sarah S. , Hortos Kari , Kovan Bruce , McCaskey Sarah , Pathak Dorothy R. , Fenton Jenifer I.
Published: June 2021

Obesity is known to increase the risk of colon cancer, and most colorectal cancers develop from adenomatous polyps. However, the effects of obesity on colonic polyp formation are unknown. The aim of this study was to determine if BMI was associated with advanced colon polyps in males. 126 men (48 – 65 yr) were recruited at time of first colonoscopy, and anthropometric measures were collected. Odds ratios were determined using logistic regression for polyp number (0 or ≥ 3) and polytomous logistic regression for polyp severity (no polyp, hyperplastic polyp, tubular adenoma). 41% of the men in our study were obese (BMI ≥ 30). Obese participants were 6.8 (CI: 1.4 – 34.6) times more likely than those who were lean (BMI < 25) to have ≥ 3 polyps than no polyps. For each category increase in BMI (lean, overweight, obese), an individual was 2.5 (CI: 1.2 – 5.4) times more likely to have ≥ 3 polyps than no polyps. Additionally, relative to lean individuals, obese individuals were 7.7 (CI: 2.0 – 29.7) times more likely to have a tubular adenoma then no polyp. As BMI category increased, participants were 2.8 (CI: 1.5 – 5.2) times more likely to have a tubular adenoma than no polyps. Due to the increased number and severity of polyps, earlier screening for polyps in obese individuals may be warranted.

Grant Funding Source: NCI 1R03CA142000 and MSU CTSI
Regulation of the Na+,HCO3‐ cotransporter NBCn1 (SLC4A7) by a constitutively active ErbB2 receptor in MCF‐7 breast cancer cells
Authors: Gorbatenko Andrej , Olesen Christina , Valen Eivind , Pedersen Stine Falsig
Published: June 2021

Net acid production is greatly increased in tumors due to a shift towards glycolytic metabolism. We previously showed that the Na+,HCO3− cotransporter NBCn1 (SLC4A7) is strongly upregulated in MCF‐7 breast cancer cells upon expression of a truncated, constitutively active ErbB2 receptor, ΔNErbB2. Here, we explore the mechanisms involved in this upregulation. NBCn1 mRNA and protein levels were reduced by inhibition of ERK or Src in ΔNErbB2‐, but not vector‐expressing MCF‐7 cells. PI3K‐Akt inhibition reduced NBCn1 expression in both cell types. NBCn1 expression was increased by stimulation of full‐length ErbB receptors. In luciferase promoter assays, the 1.2 kb genomic region upstream of SLC4A7 increased luciferase activity after ΔNErbB2 expression ~2.5 fold. Promoter deletions identified the ΔNErbB2‐ responsive region and putative transcription factor binding sites. NBCn1 expression was increased by expression of dominant‐negative (DN)‐Sp1 or by Sp1 knockdown, and decreased by knockdown of Krüppel‐like factor 4 (KLF4), while DN‐Sp3 or Sp3 knockdown had no effect. Chromatin immunoprecipitation assay (ChIP) demonstrated Sp1, Sp3 and KLF4 binding to the NBCn1 promoter. In conclusion, we identified an NBCn1 promoter and demonstrated that NBCn1 transcriptional regulation involves Sp1 and KLF4, and that ErbB2‐dependent upregulation of NBCn1 is controlled at least in part by ERK‐ and Src signaling.

Funding: Danish Cancer Society, Danish Research Council
Francisella bacteria encode genes required for efficient invasion and intracellular replication in hepatocytes
Authors: Law HT , Sriram Aarati , Fevang Charlotte , Guttman Julian Andrew
Published: June 2021

Francisella tularensis is a bacterium that can cause up to 60% mortality in humans. These microbes carry a cluster of conserved genes present in all F. tularensis subspecies called the Francisella pathogenicity island (FPI). Within the FPI, iglB, iglC, pdpA and vgrG genes are important for intracellular macrophage survival and virulence; yet, their role in epithelial cell infections, a major contributor to the progression of disease, is unclear. Given that F. tularensis uses dissimilar mechanisms to enter different cell types, we hypothesized that FPI genes play a crucial role in bacterial entry into epithelial cells. To test this hypothesis, BNL CL.2 hepatocytes were infected with F. novicida for up to 48 h. Detailed microscopic examination and gentamicin protection assay revealed that ΔiglB, ΔiglC, ΔvgrG, ΔpdpA Francisella mutants were invasion and replication deficient. We showed that gene complementation restored intracellular growth. Our data suggest that IglB, IglC, PdpA and VgrG are important for hepatocyte infections, as they are vital for efficient invasion and intracellular proliferation.

Grant Funding Source: NSERC, CIHR
Functional Characterization of the Notch Response Element: Mastermind selectivity and interaction with the TATA box determines activation strength
Authors: Ramos Cassandra Agbayani , Burke Thomas Michael , White J. Brandon
Published: June 2021

Notch signaling plays a crucial role in cell development and proliferation. The Notch receptor is an inducible transcription factor found on the cell surface which is cleaved resulting in the release of the intracellular domain (NICD). The NICD enters the nucleus and complexes with CBF1 and Mastermind (MAM) to activate target genes. Recent studies have shown that a paired CBF1 binding site is important for target gene activation. There are four potential CBF1 binding sites in Hes1 and up to six sites in Hes5. We have undertaken a series of experiments to determine what is required to activate target genes. Site‐directed mutagenesis was used to mutate the different CBF1 binding sites in the Hes1 and Hes5 promoters, change the spacing and orientation between sites, as well as mutate the TATA box and the spacing between the TATA box and the CBF1 sites. Our results confirm previous results indicating that the paired CBF1 sites are the most important in activation of target genes. We also show that the unpaired sites do not contribute significantly to target gene activation. Finally, our results indicate that the ternary Notch complex requires a TATA box to activate target genes. Mutation of the TATA box decreased reporter gene activity to basal levels. Our results suggest that an SPS and TATA are necessary and sufficient to activate Notch responsive genes. Support of research was through NSF‐RUI Grant#1052039 to JBW.
Development of a mass spectrometry assay to measure ribonucleotide reductase activity in human cancer cells
Authors: Tong Ai Phuong S. , Rice Kevin P
Published: June 2021

Ribonucleotide reductase (RNR) catalyzes the reduction of ribonucleotides into deoxyribonucleotides, which are essential for DNA replication and cell division. RNR is especially abundant in proliferating cancer cells and is an increasingly popular target in anticancer research. We had previously reported that the experimental anticancer prodrug Laromustine inhibits the enzyme thioredoxin reductase (TrxR), which is involved in proper RNR activity. Laromustine has been used in clinical trials to treat patients with acute myelogenous leukemia and glioblastoma multiforme. In this study, we use a particular sample preparation method and HPLC coupled to high‐resolution time of flight mass spectrometry to quantify deoxyribonucledotide diphosphate pools in human cells. These experiments will allow us to assess changes in RNR activity from cultured human cancer cells in response to treatment with Laromustine. We hope to further elucidate the mechanism by which Laromustine kills cancer cells, possibly influencing the clinical applications of Laromustine or similar agents.
Gold Nanoparticles Associated with Neutral Red as potential drug delivery for photodynamic therapy
Authors: Santos Naiara T , Verissimo Tanira V , Silva Jaqueline R , Azevedo Ricardo B , Gomes Anderson J , Lunardi Claure N
Published: June 2021

Gold Nanoparticles (AuNP‐TG) have been synthesized with surface modification to improve the interaction to neutral red (NR). We used AuNp‐TG NR system to evaluate possible clinical application in drug delivery system in photodynamic therapy (PDT)using visible light irradiation producing lethal agents that inactivate tumor cells. AuNP‐TG was prepared by sodium citrate reduction method. The interaction of neutral red with AUNP‐TG was performed adding increased volumes of neutral red to AuNP‐TG solution. NanoZetasizer equipment (Malvern Instrument) was used for recording the zeta potential and size distribution measurement. The AuNP and AuNP‐TG loaded with NR were evaluated in term of toxicity in NIH‐3T3 fibroblast and 4T1 tumor cells line. The distribution analyses (DLS) show a uniform size with varying diameters of AuNP‐TG (60,3nm); AuNPTG‐NR (165nm) systems. Also was estimated the number of surface atoms, Ns2 of NR on a single AuNP and AuNP‐TGA. The compound NR and AuNP‐TG‐NR induced toxicity in 4T1 tumor cell rather than NIH‐3T3 fibroblast under visible light irradiation. The modification on the surface of AuNP with TGA prevent the nanoparticle aggregation. The ability of carry more NR can be an advantage as drug delivery systems for PDT.CNPq, FAPDF,DPPUnB, FINATEC
Effects of tributyltin exposures on secretion of Interleukin 6 from human immune cells
Authors: Martin Tyesha C , Whalen Margaret M.
Published: June 2021

Tributyltin (TBT) is an organotin compound that contaminates the environment and has been found in human blood samples. TBT‐exposed mammals have shown increased incidences of tumors. Previously, we have shown that TBT was able to interfere with the ability of human natural killer (NK) lymphocytes to lyse tumor target cells. Our recent studies have shown that TBT is able to alter the secretion of the pro‐inflammatory cytokine tumor necrosis factor alpha (TNFα) from immune cells. The results indicate that concentrations of TBT of 200 – 2.5 nM caused significant alterations of TNFα secretion. Interleukin 6 (IL‐6) is produced by T lymphocytes and monocytes and is important in regulating our immune response to infections and tissue damage. Due to the important role that IL‐6 plays in immune responsiveness, it is important to understand whether exposures to TBT are able to disrupt its secretion. Peripheral blood mononuclear cells (PBMCs) were exposed to TBT at concentrations of 0–200nM and monitored for secretion of IL‐6 at 24h, 48 h, and 6 days using enzyme‐linked immunosorbent assay (ELISA). TBT at concentrations of 10–200 nM for 24 h, 48 h, or 6 days decreased the levels of IL‐6 secreted by PBMCs. Supported by NIH grants 2T34GM007663–3 and 5U54CA163066–02.
The effects of recall sequence/mode and day of week on the estimation of usual intake from 24‐hour recall data: An analysis of the 2003–2004 National Health and Nutrition Examination Survey
Authors: Kirkpatrick Sharon I , Guenther Patricia M , Dodd Kevin W
Published: June 2021

The concept of usual intake is salient to monitoring the nutritional status of populations and to assessing relationships between diet and disease. In estimating means and distributions of usual intake using 24‐hour recall data, it may be important to consider factors such as recall sequence, mode of data collection, and day of week. Using data from the 2003–2004 National Health and Nutrition Examination Survey (n = 7647), we examined the influence of recall sequence and mode (first recall, completed in person versus second recall, completed by phone) and day of week (weekday versus weekend) on mean intakes of energy, food groups, sources of empty calories, and a number of nutrients of public health significance. Although the pattern of effects differed by dietary component and among age/sex subgroups, both sequence/mode and day of week had significant effects on mean intakes. In some cases, the differences appear large enough to be of practical significance. For example, 19–70 year olds reported 100 fewer calories and 2.5 fewer teaspoons of added sugars on average on the second recall compared to the first. Sequence/mode and day of week effects also appear large enough to impact estimated usual intake distributions. The findings highlight the need to consider potential nuisance effects when modeling usual intake.
Facebook successfully recruited low‐income participants to nutrition program evaluation
Authors: Wamboldt Patricia , Lohse Barbara
Published: June 2021

A Facebook impression (ad) was tested as a recruitment strategy for evaluation of digitally‐delivered nutrition programs about family meals (FM) and folic acid (FA). Ads targeted English‐speaking, low‐income PA residents 18 – 55 y with key profile words (e.g., SNAP, food bank). Ads linked to pre and post program evaluation surveys developed with Qualtrics Pro (Provo, UT). Ad clicks were recorded for 807 FM and 795 FA responders; surveys were initiated by 73 and 109, respectively. Completers (n=47 FM; 70 FA) were similar in demographics, cognitive behaviors, and evaluative effort. FM completers were white (75%), young (mean age 35.5 ± 9.4 y), overweight/obese (59%; mean BMI 30.0 ±7.) and 51% were low‐income with 22% SNAP participation. FA completers were white (69%), young (mean age 36.1 ± 11.9 y), overweight/obese (63%; mean BMI 29.6 ±8.6,) and 51% were low‐income with 28% SNAP participation. Evaluation findings informed program revision decisions. Number of ad clicks determined overall costs for FM and FA, which were $1197.45 and $1321.52 respectively. Respective FM and FA evaluation recruitment costs were $25.48 and $18.88/person; recruitment cost for each low‐income evaluator was $39.92 (FM) and $27.53 (FA). Advantages of cost, efficiency, and speed associated with Facebook were moderated by duplicated response incidents, which were detected and considered in analyses and for future use. USDA funded.