bluuebunny/crossref_metadata_2025 huggingface dataset:
Apigenin Modulation of Whole Transcriptome Patterns of TNFα ‐ Induced Changes in Triple Negative Breast Cancer Cells, MDA‐MB‐231
Authors: Bauer David , Mazzio Elizabeth , Soliman Karam F.
Published: November 2023

Mortality associated with breast cancer is primarily attributable to metastasis, of which TNFα plays a central orchestrating role. TNFα acts on the breast tumor TNF receptors evoking the release of chemotactic proteins (e.g. MCP‐1/CCL2). These proteins directs massive inward infiltration/migration of tumor‐associated macrophages (TAMs), tumor‐associated neutrophils (TANs), myeloid‐derived suppressor cells (MDSCs), T‐regulatory cells (Tregs), T helper IL‐17‐producing cells (Th17s), metastasis‐associated macrophages (MAMs) and cancer‐associated fibroblasts (CAFs). These tumor embedded infiltrates collectively enable immune evasion, tumor growth, angiogenesis, and metastasis. In the current study, we investigated the potential of apigenin, a known anti‐inflammatory constituent of parsley, to alter the transcriptome patterns in TNFα treated triple negative breast cancer cells (MDA‐MB‐231). Whole genome expression profiling was carried out on Agilent Human 4 × 44 k arrays. The findings show TNFα mediated upregulation and apigenin downregulation of transcription patterns for the following genes: LAMC2, TMEM132A, TNFAIIP2, ICAM1, TLR2, IL32, IL4l1, CTSS, NEURL3, GBP5, IDO1, C3, EB13, ANO09, C1QTNF1‐AS1, BMPER, TNFSF15, IL 24, LINC00996, TRA2A, MAP3K5, LRG1, NEDD4l and PDA15. Pathway analysis shows the primary pathway involved in the effects of TNF were in the cytokine‐cytokine receptor interaction pathway, with sub‐categorical targets within the family of chemokines, TNF family and TGF‐Beta families. These data provide a general transcriptome pattern changes influence by TNFα, and modulated by apigenin in human breast cancer cells

Support or Funding Information

This research was supported by the National Institute of Minority Health and Health Disparities of the National Institutes of Health through Grant Number G12 MD007582 and Grant Number P20 MD006738.
Gradient Irradiation Enhances Cell Killing Efficiency via Bystander Effects in MCF‐7 cells
Authors: Zuo Li , Zhou Tingyang
Published: November 2023

In recent years, a large body of evidence has shown that irradiated cells can produce signaling factors to reduce the survival of neighboring less‐ or non‐irradiated cells, termed as bystander effects. Bystander effects may play an important role in non‐uniform irradiation through the enhancement of cell killing efficacy at the under‐dosed regions. In this study, we tested the hypothesis that a gradient dose design may achieve better tumor suppression compared to uniform‐dose distribution in breast cancer cells (MCF‐7). Cells were divided into three groups receiving 8‐2 Gy (gradient irradiation, GI), 5 Gy (uniform irradiation, UI) or 0 Gy (control), respectively, followed by viability and apoptosis assays at 2, 24, and 48h after irradiation. Data were analyzed using one‐way ANOVA in SPSS. P < 0.05 was considered statistically different. Our results show that both GI and UI caused significant reduction of cell survival and increased apoptosis at 2, 24 and 48h as compared to control (p < 0.05). Although at 2h the lower‐dose irradiated cells under GI showed higher viability and lower apoptosis than those of the cells receiving higher doses, the survival difference across the various regions of GI completely diminished at 24h and 48h. More cell death and apoptosis were observed in different areas of GI field as compared to those of the UI at 24 and 48h after irradiation (p < 0.05). This study highlights a superior therapeutic effect of GI over UI due to the bystander signaling.

Support or Funding Information

American Physiological Society (APS) 2016 S&R Foundation Ryuji Ueno Award
Exploring the Relationship Between Kaiso and C‐Terminal Binding Protein in Breast Cancer in Women of African Ancestry
Authors: Caban‐Ureña Ambar Beatriz , Jones Alana , Shin Jeehye , Park Sam , Gil‐Hernández Sara , Yan Tingfen , Kabbout Mohamed , Liang Genqing , Byun Jung , Gardner Kevin
Published: November 2023

Breast Cancer is a heterogeneous disease at the molecular and clinicopathological levels and it is well known to be influenced by internal and external factors. Therefore, this study intends to explore the relationship of two important transcription factors that have been observed in several types of cancer: C‐Terminal Binding Protein (CtBP) and Kaiso. CtBP is a transcriptional repressor that has been shown to interact with a wide range of proteins and act as a metabolic sensor through NAD+/NADH levels. On the other hand, Kaiso is a member of the POZ/ZF family that can bind to the methylated CpG dinucleotides or to the non‐methylated sequence, known as the Kaiso binding site (KBS). In previous studies, both transcriptional factors have demonstrated a worse prognosis and survival in cancer patients. Hence, our objectives are determining the relationship and possible molecular links behind these two proteins, by focusing on the transforming growth factor‐β (TGFβ) signaling; an essential pathway in the epithelial to mesenchymal transition (EMT). To accomplish our goal, we have been working with a sample size of 559 breast cancer patients in which we have collected both patient and tumor characteristics and parameters. They include: BMI, adipocyte cross‐sectional area, race, ethnicity, pre or post‐menopausal status, age and mitotic rate. In addition, immunohistochemistry (IHC) was performed using tissue microarrays for: ER, PR, Kaiso, CtBP, pSmad and other molecules associated with TGFβ signaling. Statistical analysis was performed to observe the correlation between the proteins and patient outcome, based on hormone status, race and the activation of the TGFβ signaling pathway. Preliminary results indicate a relationship between Kaiso, CtBP and survival that is modified by race. Further studies are directed at defining the mechanism underlying this relationship.
Actual and Potential Use of Information Technology in Pharmacology Education Among Medical Students in a Developing Country
Authors: Kwanashie Helen Ochuko
Published: November 2023

Information Technologies (IT) have become highly acclaimed tools in teaching‐cum‐learning generally, including in pharmacology education. A survey was carried out among male and female clinical medical students aged approximately 24 years in northern Nigeria, in an attempt to determine their actual and potential use of IT in learning pharmacology. Over 90% of the students owned personal laptops/notebook computers and smartphones which were used by them primarily for educational (88%) and leisure/entertainment (75%) purposes. Educational use involved an average of 40% of their private study time (range being 10–70%), and included receiving resources from lecturers in addition to exchange of materials with fellow students. Internet platforms mostly used were whatsapp (100%), email (94%), facebook (94%) and google (94%), with many judging their internet skills to be 50–88% good. Top impediments were identified as high internet subscription costs (100%) and frequent power outages (75%). All the respondents believed that IT tools will enhance teaching‐cum‐learning of pharmacology as well as other subjects in the medical curriculum. Projected slides/printed slide‐handouts were the preferred mode to be employed by their lecturers for teaching pharmacology by 56% of the students while 44% opted for a hybrid with the more traditional mode using written or typed notes/white boards. Their suggested potential use included e‐books, video lectures, access to online libraries and interactive students‐lecturers fora. The questionnaire data was affirmed by a key informant interview of the student class representative. It is concluded that the actual and especially, potential use of IT in pharmacology education, is huge among the students studied and possibly in similar populations within and outside Nigeria ‐ requiring reforms, creativity and innovations among all stakeholders.
Funding success following STEM teacher mentoring drives classroom innovation
Authors: Rockhold Rob , Sullivan Donna , Bender Susan
Published: November 2023

From 1994 through 2013, mentoring of high school science, technology, engineering and math (STEM) teachers in grant‐writing exercises by medical center faculty resulted in submission of 158 applications for funding of inquiry‐based classroom activities, of which 123 (78%) were funded and which brought over $575,000 of new funds into classrooms. The Base Pair Summer Research Institute at the University of Mississippi Medical Center, funded by the Howard Hughes Medical Institute, provided directed training in identifying funding sources, describing needs and delineating defined outcomes to 157 high school STEM teachers for periods of 1 to 6 weeks. Of these, 37 (24%) responded to a 2016 online survey, 31 of whom had submitted at least one application (mean of 3.6 grants/teacher, range 1–7). The majority (13) garnered from $500–$2,000 from funded applications, but 5 teachers each gained a total of over $10,000. Teachers (77%) receiving grants expressed “much more” or “more” job satisfaction, while 74% felt grant writing increased a perception of professionalism, and 83% indicated that the funding “greatly increased” or “increased” their ability to teach effectively. Students (93%) were reported to be “much more engaged” or “more engaged” in the classrooms using grant funds. Despite these findings, only 30% of teachers reported receiving “much more” (2) or “more” (7) recognition from their school administration for receipt of funding. Mentoring of high school teachers by medical center faculty builds professionalism and job satisfaction among teachers, bolsters student engagement, and provides critically needed monetary support for educational innovation in STEM classrooms. Increased efforts are needed to address the inconsistent recognition of such professional development by secondary school administrators.

Support or Funding Information

(Supported by Precollege Science Education awards from the Howard Hughes Medical Institute.)
The Effects of THC on Brain Mitochondria of Zebrafish
Authors: Awodu Olamide , King Adrienne Lester
Published: November 2023

Tetrahydrocannabinol (THC) is a cannabis and is the primary psychoactive component of marijuana. Cannabinoids can exert various biological effects. It has been shown to have therapeutic applications in the medical field such as treating medical pathologies: pain, epilepsy, and movement disorders. However, there are additional studies showing the negative impacts of THC. In particular, cannabis might directly affect brain function with a special emphasis on mitochondrial function. Therefore, we hypothesize that THC will decrease mitochondrial function. The aim of this study was to analyze the effects of THC on brain mitochondrial function using the model organism Danio.rerio (zebrafish).

Methods

Zebrafish were exposed to 20 mg/ml of THC and brain mitochondria was isolated. Oxygen consumption in the presence of glutamate/malate was monitored to determine mitochondrial function, and results compared to control zebrafish. Basal mitochondrial respiration (State 3 and State 4) was measured using a Clark‐type oxygraph electrode system. The respiratory control ratio (RCR) was also calculated by taking the ratio of state 3 divided state 4.

Results

Zebrafish exposed to THC had a significantly lower State 3 respiration compared to control (n=6, p <0.05). A decrease in state 3 respiration is notable because it indicates that THC decreases electron transport and subsequently the rate at which brain mitochondria may synthesize ATP. Moreover, the RCR was significantly lower in THC treated fish (n=6 p<0.05). A higher RCR indicates mitochondria that are more tightly coupled with better function (i.e., better quality of mitochondria), whereas a lower RCR indicates that mitochondria are more loosely coupled with poorer function (i.e., damaged mitochondria) in response to treatment.

Conclusion

These data demonstrate that the bioenergetics of brain mitochondria are impaired and THC does appear to have an impact on mitochondrial function.
Persistent Potentiation of the Analgesic Effects of Opioids by Delta‐9Tetrahydrocannabinol (THC) in Nonhuman Primates
Authors: Nilges Mark R. , Winsauer Peter
Published: November 2023

Improving the analgesic effectiveness of opioids while also reducing the adverse effects is a major goal of pain research. One approach to this problem has been to combine drugs such as cannabinoids with opioids to determine if this might reduce the dose of opioid required to produce analgesia. In this experiment, male nonhuman primates (rhesus macaques) were administered a range of doses of THC in combination with cumulative doses of heroin to determine if the combinations produced greater analgesia than that observed with heroin alone. Analgesia was assessed using the tail‐dip procedure. Briefly, the animals were secured in standard primate chairs, tails were shaved, and dipped in a mixed order in three different water temperatures (40 °, 50 °, and 55 °C) until the animal rapidly withdrew his tail, or 20 seconds elapsed. Therefore, latency (sec) to remove the tail was converted to % maximum effect (%MPE). Each set of tail dips was separated by a 15‐min period that also served as an injection interval for the cumulative doses of drug. The initial tail dips of a session served as the baseline, whereas the initial injections were either vehicle (control) or THC (0.056–0.18mg/kg, i.m.). Sessions usually occurred once per week, and occasionally, twice per week. Prior to testing THC and heroin in combination, buprenorphine (n=7), morphine (n=4), and heroin (n=7) were administered alone (0.032–3.2 mg/kg, i.m.) for comparison purposes. These same opioid agonists were also administered after the THC‐heroin combinations. At the doses of each drug tested, the rank order of analgesic effectiveness was heroin>buprenorphine>morphine, with maximum %MPE differing between the two high temperatures. For 50°C, the %MPE were 60%, 29%, and 27%, respectively. For 55°C, %MPE were 42%, 16%, and 7%, respectively. When THC was administered prior to heroin, there was a similar one‐half log leftward shift in the heroin dose‐effect curve produced by all three doses of THC. Unexpectedly, the leftward shift in the heroin dose‐effect curve was still evident in the absence of THC when heroin alone was re‐determined after a drug‐free period of approximately 1 week. These effects were mediated by mu opioid receptors since naltrexone (0.1mg/kg) reversed the analgesic effects of heroin at both water temperatures. A leftward shift in the morphine dose‐effect curve was also evident 2–3 weeks after the THC‐heroin combinations. Unlike heroin and morphine, the dose‐effect curve for buprenorphine was not shifted relative to the initial curve; however, it was tested almost 2 months after the THC‐heroin combinations. In conclusion, acute doses of THC administered as a pretreatment potentiated the analgesic effects of heroin acutely, and for an extended period after the administration of the combination.
Antinociceptive Effects of Novel GABAA Receptor Positive Allosteric Modulators in Mice: Pain‐Stimulated and Pain‐Depressed Behaviors
Authors: Lewter Lakeisha , Cook James M , Li Jun‐Xu
Published: November 2023

Preclinical pain and analgesia research has primarily focused on pain‐stimulated behaviors, despite the fact that pain often depresses certain behaviors. Positive allosteric modulators (PAMs) that are selective for α2/α3 subunit‐containing GABAA receptors have been shown to mediate profound analgesia in animal models of inflammatory and neuropathic pain. However, the ability of α2/α3 GABAA receptor PAMs to restore pain‐depressed behaviors have not been explored. This study sought to examine the effects of two novel α2/α3 selective PAMs, KRM‐II‐18B and KRM‐II‐81, on acetic and lactic acid‐induced writhing (pain‐stimulated behavior), and two pain‐depressed behaviors: acid‐induced suppression of locomotor activity and acid‐induced depression of nesting behavior in mice. Drug effects of PAMs were compared with currently available analgesics morphine and acetaminophen. Intraperitoneal (i.p.) administration of acetic acid (0.6%) and lactic acid (0.32%) induced stimulation of writhing, depression of locomotor activity, and depression of nesting behavior. Morphine, KRM‐II‐18B, and KRM‐II‐81 pretreatment all produced dose‐dependent and significant decreases in both acetic acid and lactic acid‐induced writhing. This effect was attenuated by the benzodiazepine receptor antagonist flumazenil, confirming that the analgesic effects of KRM‐II‐18B and KRM‐II‐81 are mediated through the benzodiazepine binding site of GABAA receptors. While pretreatment of acetaminophen was able to block acid‐depressed locomotion, pretreatment of both PAMs failed to block acid‐depressed locomotor activity. In the nesting procedure, morphine, KRM‐II‐18B, and KRM‐II‐81 failed to alleviate acetic acid‐induced depression of nesting behavior, but dose‐dependently attenuated lactic acid‐induced depression of nesting. Taken together, these findings suggest that the ability of subunit‐selective PAMs to alleviate pain‐depressed behavior is moderately assay dependent and also depends on the degree of pain.

Support or Funding Information

This research was supported by RO1DA034806.
Effects of α7 nicotinic receptor positive allosteric modulator on sodium‐potassium‐chloride co‐transporter 1 activation following lipopolysaccharide‐induced neuroinflammatory pain in mice
Authors: Rahman Shafiqur , Abbas Muzaffar
Published: November 2023

Neuroinflammatory pain is a chronic neurological disorder characterized by allodynia and hyperalgesia. We have shown that 3a,4,5,9b‐tetrahydro‐4‐(1‐naphthalenyl)‐3H‐cyclopentan[c]quinoline‐8‐sulfonamide (TQS), an α7 nicotinic acetylcholine receptors (α7 nAChRs) positive allosteric modulator (PAM), reduces neuroinflammatory pain, and microglial activation in the hippocampus in mice. The objective of the present study was to determine the effects of TQS, and bumetanide (BU), sodium‐potassium‐chloride co‐transporter 1 (NKCC1) antagonist, on lipopolysaccharide (LPS)‐induced neuroinflammatory pain using von Frey hair test, a widely used method for the quantification of tactile allodynia and hot plate test, a widely used technique for thermal hyperalgesia assessment, in male mice. In addition, we examined the effects of TQS (4 mg/kg) on NKCC1 mRNA expression and p‐NKCC1 immunofluorescence in the hippocampus. TQS (4 mg/kg) significantly decreased LPS‐induced tactile allodynia and thermal hyperalgesia. In addition, co‐administration of BU (10 mg/kg) with TQS (1 mg/kg) reduced tactile allodynia and thermal hyperalgesia. Taken together, these results suggest that TQS decreases LPS‐induced neuroinflammatory pain by reducing NKCC1 activation through modulating microglial α7 nAChR allosteric site.

Support or Funding Information

Supported by Fulbright Foundation USA
Lasting anti‐hyperalgesic effect of nanoemulsion containing essential oil of H. pectinata in an animal model of fibromyalgia
Authors: Quintans Lucindo Jose , Quinans Jullyana Siqueira , Camargo Zaine , Arrigoni‐Blank Maria F , Brito Renan , Oliveira Marlange , Santos Marcio , Araújo Adriano , Kolker Sandra J , Sluka Kathleen
Published: November 2023

Hyptis species are aromatic plants that produce essential oils which present pharmacologically active substances with antimicrobial, anti‐HIV, analgesic and antiinflammatory activities. We previously demonstrated the analgesic profile of leaf essential oil of Hyptis pectinata, a medicinal plant widely used in Brazilian folk medicine, however, its pharmacological effect is not durable enough for possible applicability in chronic disorders. Thus, we developed and evaluated a nanoemulsion containing leaf essential oil of H. pectinata (NE‐EOH) produces a long‐lasting anti‐hyperalgesic effect on chronic musculoskeletal pain animal model. The NE‐EOH was prepared by addition of 20 wt% Pluronic F127 and showed hydrodynamic diameter and zeta potentials of 220±28 and 10.1±7.2, respectively. Mice were assessed for mechanical hyperalgesia of the paw and muscle (von Frey and tweezer), motor coordination (Rota Rod) and muscle strength (Grip Strength Meter) beginning 1 h after treatment and continuing through 11 days. NE‐EOH, one single treatment (first day only), produced a significant reduction (p<0.05 or p<0.01) of hyperalgesic behavior. This analgesic activity was reversed by both naloxone and methysergide. Moreover, NE‐EOH reduced (p<0.05) substance P levels in neurons from the spinal cord. Thus, NE‐EOH produced a lasting antihyperalgesic effect probably mediated by brainstem inhibitory pathways or reducing excitatory neurotransmitters in the spinal cord.

Support or Funding Information

CNPq/Brazil [Grants # 441969/2014‐3, 200636/2012‐0 and 470774/2011‐8], FAPITEC/SE/Brazil, CAPES/Brazil and FINEP/Brazil.
D‐Limonene complexed with β‐cyclodextrin reduces hyperalgesia in a mice model for fibromyalgia
Authors: Barreto Rosana SS , Lima Pollyana SS , Araujo Heitor G , Pereira Erik WM , Rezende Marilia M , Guimaraes Adriana G , Barreto Andre S , Santos Marcio RV , Araujo Adriano AS , Quintans‐Junior Lucindo J , Quintans Jullyana SS
Published: November 2023

D‐limonene (LIM) is a monocyclic monoterpene with a lemon‐like odor and is a major constituent in several citrus oils (orange, lemon, mandarin, lime, and grapefruit). The analgesic profile of LIM has been described previously. However, the short half‐life and water insolubility has been a limitation to LIM medicinal uses. So, cyclodextrins, such as β‐cyclodextrin (β‐CD), has been an important tool to minimize these limitations of non‐polar compouns, as LIM. Now, we assessed the anti‐hyperalgesic effect of LIM‐β‐CD complexed or LIM alone in an animal model of chronic musculoskeletal pain. Thus, mice model of musculoskeletal pain was performed as reported by Sluka et al. After we confirmed hyperalgesic behavior, mice were treated with LIM‐β‐CD or LIM (50 mg/kg; p.o.) or vehicle (saline 0.9%, p.o.) or tramadol (TR, 4 mg/kg, i.p.). The mice treated with LIM‐β‐CD showed significant (p<0.001) superior anti‐hyperalgesic effect when compared with LIM alone, these effects can not produced any alteration in the grip strength force. Additionally, we perfused the lumbar spinal cord collected, crioprotected, cut and submitted in an imunofluorescence protocol for Fos protein. Both, LIM‐β‐CD or LIM, significantly (p<0.001) reduced expression of Fos protein on spinal cord horn. Thus, we can suggest that β‐CD improves the anti‐hyperalgesic effect produced by LIM and this effect seems to be due the involvement of descending pain inhibitory mechanisms.

Support or Funding Information

CNPq, CAPES, FAPITEC‐SE and FINEP/Brazil
TRPV1 Mediates The Orofacial Antinociceptive Effect of Frutalin: In Vivo and In Silico Studies
Authors: Barros Adriana Rolim Campos , de Barros Mamede Vidal Damasceno Marina , Santos Sacha Aubrey Alves Rodrigues , de Maria de Albuquerque Melo José , Magalhães Francisco Ernani Alves , Vieira‐Neto Antonio Eufrásio , de Azevedo Moreira Renato , de Oliveria Monteiro‐Moreira Ana Cristina
Published: November 2023

Our previous studies have shown the orofacial antinociceptive effect of frutalin (FTL) and it was suggested that FTL effect on acute orofacial pain could be mediated by TRPA1 and TRPV1. Docking study indicated that FTL strongly interacts with TRPV1. This study was conducted to investigate the relationship between TRPV1/TRPA1 channels and the orofacial antinociceptive effect of FTL. Mice (n=6/group) were pretreated with vehicle (0.9 % NaCl; 10 mL/kg; i.p.), FTL (0.5 mg/kg; i.p.), ruthenium red (TRPV1 antagonist, 3 mg/kg, s.c.) or camphor (TRPA1 antagonist, 7.6 mg/kg, s.c.) 30 min before injection (20 μL) of capsaicin (TRPV1 agonist, 1.6 μg/lip) or cinnamaldehyde (TRPA1 agonist, 13.2 μg/lip). In order to verify the involvement of TRPV1 and TRPA1 channels, others groups of mice were pretreated with the respective antagonist 15 min before administration of FTL. Nociception was quantified as time (s) spent rubbing the site of injection 0–5 min after induction. The interaction between FTL and TRPA1 was analyzed using molecular docking. The HEX software was used for molecular docking calculations and the direct action of FTL on TRPA1 was viewed using PyMOL v1.4.7. Experimental protocols were approved by the UNIFOR Animal Research Ethics Committee (#0122014). Pretreatment with FTL was associated with a reduction in the intensity of face rubbing induced by capsaicin (***p<0.001) or cinnamaldehyde (****p<0.0001). Ruthenium red, but not camphor, abolished the antinociceptive effect of FTL. It was possible to observe a direct action of FTL amino acids (Asp149, Tyr146, Trp 147 and Gly25) on TRPA1 glycosylated amino acids (1.8–3.6 Å). However, out of 10 more energetic clusters, only 4 overlapped, suggesting that there are repulsions and chemical impediments that hinder the FTL fitting more than once in the same site of the channel. The energies observed in the 10 investigated clusters are high, but do not exceed those previously observed for TRPV1 channel. FTL antinociceptive effect was abolished by the TRPV1 antagonist, but not by the TRPA1 antagonist and FTL. Besides this, FTL strongly interacts with TRPV1 and it has a weak interaction with TRPA1. Since and TRPV1 and TRPA1 channels are co‐expressed and they interact in neurons, we suggest that FTL effect is mediated by TRPV1.

Support or Funding Information

Edson Queiroz Foundation (FEQ), Cnpq and Capes.
Ca2+ Signaling Inhibitors Attenuate the Antinociceptive but not Discriminative Stimulus Effects of the Imidazoline I2 Receptor Agonist 2‐BFI in Rats
Authors: Siemian Justin Nicholas , Zhang Yanan , Li Jun‐Xu
Published: November 2023

Recent research has established the imidazoline I2 receptor as a promising target for the development of novel analgesics. However, despite a strong understanding of imidazoline I2 receptor‐mediated behavioral effects, little is known about its post‐receptor signaling mechanisms. This study examined the effects of several inhibitors of Ca2+ signaling mechanisms on two behavioral effects of the prototypical imidazoline I2 ligand 2‐BFI. The von Frey test was used to examine the antinociceptive effects of 2‐BFI in complete Freund's adjuvant (CFA)‐induced inflammatory pain in rats. A two‐lever drug discrimination paradigm in which rats were trained to discriminate 5.6 mg/kg (i.p.) 2‐BFI from saline was used to examine the discriminative stimulus effects of 2‐BFI. The L‐type Ca2+ channel blockers verapamil and nimodipine, the calmodulin antagonist W‐7, and the internal Ca2+ release inhibitor ryanodine dose‐dependently attenuated the antinociceptive effects of 2‐BFI. Oxycodone‐induced antinociception was unaffected by verapamil and nimodipine. In contrast, verapamil, nimodipine, and W‐7 did not affect the discriminative stimulus effects of 2‐BFI. These results suggest that the antinociceptive effects of 2‐BFI involve intracellular Ca2+ elevation and/or downstream Ca2+/calmodulin signaling, whereas the discriminative stimulus effect of 2‐BFI is mediated by a distinct, independent mechanism.

Support or Funding Information

R01DA034806
Cocaine as Opposed to Methamphetamine Did not Restore CirWadian Activity Rhythms in Arrhythmic Suprachiasmatic Nucleus Lesioned C3H/HeN Mice when Paired with Running Wheel
Authors: Vega Juliany Yari Marrero , Clough Shannon , Dubocovich Margarita L
Published: November 2023

The “master biological clock” or suprachiasmatic nucleus (SCN) orchestrates synchrony among central and peripheral circadian oscillators. When the SCN is removed the circadian rhythm of an organism becomes arrhythmic. However, when methamphetamine (Meth), a psychostimulant, is given to mice ad libitium in drinking water, it rescues the locomotor behavioral rhythms in arrhythmic SCN lesioned (SCNX) mice (Tataroglu et al., JBiol Rhythms 2006;21;185–94). We recently demonstrated that Meth, which increases dopamine release by reversing the neuronal transporter, paired with concurrent access to running wheel (RW), is necessary for the expression of circadian entrainment in SCNX mice, possibly by activating an extra‐SCN brain oscillator (Rawashdeh et al. FASEB J, 2016). The goal of our project was to determine the ability of another psychostimulant, cocaine, which increases depolarization‐evoked dopamine release by blocking neuronal uptake, to establish rhythmic circadian activity in SCNX mice. Following the protocol described by Rawashdeh et al., 2016, C3H/HeN male mice (3–4 months old) underwent sham surgery (n=8) or SCN electrolytic lesions (n=11) (SCNX). Sham operated (free running circadian rhythms) and SCNX (arrhythmic) mice in constant dark were given a single daily vehicle (0.9% saline, i.p.) or cocaine (20 mg/kg, i.p.) injection at the beginning of an 8‐hour period of restricted RW access, for 14 days. Mice were then provided a 7‐day period of continuous access to RW to assess the development of a free running circadian activity rhythm. Our results demonstrate that SCNX mice treated for 14 days with either vehicle or cocaine paired with RW did not develop rhythmic circadian activity. This is in contrast to Meth, which when used in the same paradigm, induced a free‐running rhythm in SCNX mice (Rawashdeh et al., 2016). Although both Meth and cocaine, at doses that elicit similar levels of locomotor activity, increase the extracellular concentration of dopamine in the striatum and nucleus accumbens, the net levels of extracellular dopamine are 2–3 times fold higher following treatment with Meth (reversal of dopamine transporter) than with cocaine (blockade of neuronal uptake) (Kuczenski, et al., J Neurosci 1991;11; 2703–12). We suggest that increased level of synaptic dopamine induced by Meth paired with RW compared to cocaine may be necessary to re entrain circadian activity rhythms in arrhythmic mice.

Support or Funding Information

Supported by JSMBS funds to MLD
Evaluation of the Anticonvulsant Effects of Propofol in Pentylenetetrazole kindled Rats
Authors: Hasan Zuheir A
Published: November 2023

Propofol is a short‐acting intravenous anesthetic which is widely for the induction and maintenance of anesthesia. However, sporadic reports on the occurrence of seizures in epileptic and non‐epileptic patients, especially during recovery continue to appear. Experiments on Sprague‐Dawley rats were carried out to study the effects of propofol on clonic‐tonic convulsions in kindling phenomena evoked by multiple injections of a subconvulsive dose (40 mg/kg) pentylenetetrazol (PTZ). Kindling was considered to be completed when an animal had experienced generalized clonic–tonic seizures (stage 5). To induce complete kindling 14.0 ± 1 injections were performed. Once the animals consistently exhibit full stage of kindling, different animal groups were injected intravenously with various doses of propofol (5, 10 and 20 mg/Kg). Following the administration of propofol, kindled rats were injected with 40 mg/Kg PTZ and the seizure score was evaluated. The results of the present study showed that the highest two doses of propofol (20, 0 mg/Kg) prevent the occurrence of any kind of convulsions. The lowest dose (5 mg/Kg) did not significantly reduce the incidents of clonic‐tonic. These data suggest that propofol has marked dose dependent anticonvulsant effects on generalized clonic‐tonic convulsions in PTZ kindled rats and provides further support for the anticonvulsant effects of propofol reported in other experimental seizures models. (Supported by a grant from Arabian Gulf University).

Support or Funding Information

Arabian Gulf University
ATP Exposure Increases Expression of α‐Synuclein in SH‐SY5Y Cells: Potential Link Between Neuronal Injury and Parkinson's Disease
Authors: Lamberts Jennifer T. , Rustem Danielle , Wallis Brittnie , Zwart Colton
Published: November 2023

Parkinson's disease (PD) is a devastating neurodegenerative disorder for which there is presently no cure. The pathologic hallmark of PD is intra‐neuronal accumulation of the protein α‐synuclein (α‐syn), and in rare instances, PD has been directly linked to multiplication of the α‐syn gene locus, suggesting that overexpression of α‐syn is pathogenetic per se. It has been suggested that α‐syn aggregation is increased in cells exposed to ATP, a condition known to arise during traumatic brain injury. These results suggest a potential link between neuronal injury, purinergic signaling, and the pathogenesis of PD, yet there remains limited information about this phenomenon. The primary goal of this project was to test the effect of ATP exposure on intracellular expression and extracellular secretion of α‐syn in mammalian cells. In preliminary studies, SH‐SY5Y cells treated for 24h with 1mM or 3mM ATP expressed higher levels of α‐syn protein, as measured by Western blot. However, these concentrations of ATP also slightly reduced cell viability at 24h, as measured by CellTiter‐Blue® assay. These results further support the proposed link between purinergic signaling and PD pathogenesis. These and future studies will provide much needed insight into the role of purinergic signaling in PD and could help identify novel therapeutic targets for the treatment of PD.

Support or Funding Information

This project was funded by an internal Faculty Research Grant from Ferris State University and startup funds from the College of Pharmacy at Ferris State University.
The effect of sex on APOE‐modulated AD pathology in EFAD mice
Authors: Avila‐Munoz Maria Evangelina , Balu Deebika , York Jason , Ghura Shivesh , Collins Nicole , Ladu Mary Jo
Published: November 2023

APOE4 allele of apolipoprotein E (apoE) is the greatest genetic risk factor for Alzheimer's disease (AD) and is associated with accelerated accumulation of both amyloid plaques and and soluble oligomeric forms of the amyloid‐β peptide (oAβ), likely a proximal neurotoxin. Importantly, female APOE4 carriers have a greater lifetime risk for developing AD, an increased rate of cognitive decline and accelerated accumulation of Aβ compared to male APOE4 carriers. In vivo progress has been limited by the lack of a tractable familial AD‐transgenic (FAD‐Tg) mouse model expressing human (h)‐ rather than mouse (m)‐APOE. To study the interactions among sex, h‐APOE and AD pathology, we developed the EFAD‐Tg mice by introducing the h‐APOE genotypes into the 5×FAD‐Tg mice. We previously demonstrated that in the EFAD mice, confirmed in human control and AD samples of CSF and brain, that apoE lipidation is lower and soluble Aβ levels are higher with APOE4 vs. APOE3. Thus, we have developed the mechanistic hypothesis that AD pathology and APOE4 cause a reduction in apoE lipidation, resulting in inefficient clearance of soluble A β synaptic loss, memory/cognitive deficits, and dementia. Novel preliminary data demonstrate that the differences in female vs. male E4FAD mice mimic these established APOE4 vs. APOE3 differences in Aβ aggregation/accumulation, and apoE levels/lipidation. In addition, amyloid deposition, neuroinflammation and tau pathology is significantly greater in the EFAD females compared to males. These data suggest that sex profoundly influences APOE genotype‐specific effects on AD pathology, with apoE lipidation and oAβ levels possibly at the intersection between these two AD risk factors. Together, these data will help to address the critical need for treatment options for women at high risk for AD from the APOE4 allele.

Support or Funding Information

Current LaDu lab funding includes NIH (NIA) P01AG044682, R21AG030128, R21AG048498, R21AG051233, CCTS (UL1RR029879)
ApoE Remodeling as a Therapeutic Target for Alzheimer's Disease
Authors: Ladu Mary Jo , Avila‐Munoz Maria Evangelina , Valencia‐Olvera Ana , Ghura Shivesh , Collins Nicole , Balu Deebika , York Jason
Published: November 2023

APOE4 is the greatest genetic risk factor for Alzheimer's disease (AD), increasing risk up to 15‐fold compared to APOE3. As ɛ4 carriers can respond anomalously in clinical trials, there is a critical lack of therapeutics targeting mechanistic pathways of APOE4‐induced AD risk. APOE4 is associated with accelerated amyloid‐β (Aβ) accumulation, both as amyloid and soluble oligomeric forms of Aβ (oAβ), the latter considered a proximal neurotoxin. Using EFAD mice, a new tractable preclinical mouse model (expressing 5 familial AD (FAD) mutations and the human APOE genotypes), we have developed the mechanistic hypothesis that apoE4 is poorly lipidated compared to apoE3, resulting in lower levels of apoE/Aβ complex and higher levels of soluble oAβ. ABCA1 is a promising therapeutic target for testing this hypothesis, as it is a major transporter of lipid to nascent apoE‐containing lipoproteins in the CNS. We previously demonstrated that treatment with RXR agonists increased ABCA1 levels, apoE4 lipidation, and soluble apoE4/Aβ complex levels, while lowering soluble Aβ levels. However, while these data provide target validation for ABCA1, the detrimental effects on the liver limit RXR agonist utility in even short‐term treatment protocols. Therefore, we have identified and are evaluating alternative pharmacological approaches to enhance apoE lipidation, modeling our approach on the peripheral targets for lipoprotein biogenesis and remodeling common in the cardio‐vascular field. Two therapeutic approaches of particular interest are an ABCA1 agonist (Compound A) and an Acyl‐CoA:cholesterol acyltransferase (ACAT) inhibitor (Compound B) to increase cholesterol efflux and free cholesterol availability, respectively. In vitro, both compounds facilitated cholesterol efflux from primary glia and increased the concentration and lipidation of apoE. In vivo, both compounds are brain penetrant and further analysis is on going, including treatment and prevention paradigms with read‐outs that include biochemical and AD pathology, as well as behavioral measurements.

Support or Funding Information

P01AG044682, R21AG030128, R21AG048498, R21AG051233, CCTS (UL1RR029879)
The effect of aging on APOE‐modulated AD pathology in EFAD mice
Authors: Balu Deebika , Valencia‐Olvera Ana , Collins Nicole , Salzman Ryan , York Jason , Ladu Mary Jo
Published: November 2023

The APOE ɛ4 allele of apolipoprotein E (apoE) is the greatest genetic risk factor for Alzheimer's disease (AD) compared to APOE3, with APOE2 providing a protective effect. APOE4 is associated with accelerated amyloid‐β (Aβ) accumulation, both as amyloid plaque and soluble oligomeric forms of Aβ (oAβ), the latter considered a proximal neurotoxin. Using EFAD mice, a novel, tractable human‐APOE/familial AD‐transgenic (FAD‐Tg) preclinical mouse model, we have developed the mechanistic hypothesis that APOE4 causes a reduction in apoE lipidation, resulting in inefficient clearance of soluble Aβ, synaptic loss, and memory deficits. By 6‐months (M), E4FAD mice have greater cognitive impairment, AD pathology and soluble Aβ levels, with lower apoE lipidation levels compared to E3FAD. However, the effect of aging on EFAD mice has not been analyzed beyond 6M. In this study, at 8M, 10M, 14M and 18M, we measured amyloid deposition, neuroinflammation and tau pathology, as well apoE lipidation, and soluble levels of Aβ42, oAβ and apoE/Aβ complex. AD pathology continues with age (E4FAD > E3FAD) spreading from the subiculum and deep layers of the frontal cortex to the entire length of the cortex and particularly the thalamus. Comparison of Aβ deposition identified with an Aβ mAb vs. Thio‐S staining revealed both regional and age‐dependent changes in plaque morphology. Importantly, while the levels of soluble Aβ42 plateaued at 10M in E4FAD and 14M in E3FAD mice, oAβ levels continued to increase, from 6–18M with E4FAD > E3FAD. The predictive validity of preclinical FAD‐Tg mouse models that lack h‐APOE, the major genetic risk factor for AD, remains unclear. As the greatest risk factor for AD is age, aged EFAD mice address both these critical risk factors.

Support or Funding Information

NIH (NIA) P01AG044682, R21AG030128, R21AG048498, R21AG051233, CCTS (UL1RR029879)
(−)‐Epicatechin Mitigates Oxidative Stress, Neuroinflammation and Hyperphosphorylation of TAU in Hippocampus of Senile Mice
Authors: Ramirez‐Sanchez Israel , Rodriguez Alonso , Neri Teresa , Mendoza‐Lorenzo Patricia , Ceballos Guillermo , Villarreal Francisco
Published: November 2023

Evidence has implicated oxidative stress (OS) and inflammation as a driver of neurodegenerative pathologies such as Alzheimer's disease. We previously reported on the beneficial effects of treatment with the cacao flavanol (−)‐epicatechin (Epi) on aging‐induced OS and capacity to restore modulators of mitochondrial biogenesis in the prefrontal cortex of 26‐month‐old mice. In the present study, using a similar mouse model of aging, we examined the capacity of Epi to mitigate hippocampus OS, inflammation, hyperphosphorylation of tau protein, and soluble β‐amyloid protein levels leading to improved memory and anxiety levels. Male mice (26 month old) underwent 2 weeks of Epi treatment (1 mg/kg/day BID by oral gavage) and samples of hippocampus collected for the assessment of relevant endpoints. The assessment of the OS markers protein carbonyls and malondialdehyde levels evidence significant increases (~3 fold) with aging that are suppressed (p<0.05) by Epi. Using Western blots the protein levels of the proinflammatory molecules COX2, NFkB, TNF alpha, interleukins (1β, 5 and 6) and interferon gamma increase with aging (~2–2.5 fold) and are also significantly suppressed with treatment. Aging also leads to the hyperphosporylation of tau, increased soluble β‐amyloid levels (~ 2 fold) and reductions in nerve growth factor (~75%) which are reversed by Epi. Measures of anxiety (open field tests) and object recognition (memory) results were also improved by treatment. Altogether, results evidence a unique capacity of Epi to significantly mitigate hippocampus OS and neuroinflammation leading to improved brain function. These results may account for the beneficial effects that dark chocolate exerts on measures of memory in humans.

Support or Funding Information

NIHDK98717
Erythropoietin Reverses Cognitive Deficits Following Infantile Traumatic Brain Injury: A Translational Touchscreen Investigation
Authors: Jantzie Lauren , Robinson Shenandoah
Published: November 2023

Traumatic brain injury (TBI) is a leading cause of death and severe morbidity for infants who are born healthy. No targeted treatment currently exists to actively promote repair after early TBI, despite the high incidence of chronic, debilitating deficits. EPO is a neuro‐reparative agent with multiple mechanisms of action that may benefit infants who suffer TBI. TBI in infants increases EPO receptor expression on neural cells, and extended EPO dosing reduces neuroinflammation and enhances survival and maturation of oligodendrocytes and neurons after injury. Using a translatable preclinical model of infant TBI, we hypothesized that EPO would reverse deficits of cognition as observed on a touchscreen paradigm. Controlled cortical impact (CCI) was performed on postnatal day 12 (P12) rats of both sexes. Anesthetized rats underwent left parietal craniectomy and 0.6mm impact. On post‐injury day (pid) 1, injured rats were randomized to 6 doses of intraperitoneal erythropoietin (EPO) 3000U/kg/dose or vehicle (sterile saline) over pid 1–8. Similar to the Cambridge Neuropsychological Test Automated Battery (CANTAB) in humans, a touchscreen platform was applied to assess multiple cognitive domains. Adult rats were trained on a touchscreen operant platform by blinded investigators. Following successful training, rats performed visual discrimination (VD) and reversal tasks to assess executive function and cognitive flexibility (n=6–8/group, two way ANOVA with Bonferroni correction). Results indicate that Sham, CCI and CCI‐EPO rats are able to successfully perform VD with each group requiring a similar number of sessions to passing criteria and committing similar numbers of discrimination errors. However, when assessing reversal learning, only 57.1% of CCI animals successfully passed criteria compared to 100% of Sham and 83.3% of CCI animals treated with EPO. Notably, CCI animals committed significantly more errors and required more correction trials (1114±132) compared to Sham (730±95) and CCI‐EPO (652±61, p<0.05 for all), concomitant with increased perseveration (all p<0.05). Taken together, EPO treatment ameliorates deficits from TBI on a touchscreen platform, including error reduction and perseveration, and restores cognitive flexibility. EPO, administered in an extended post‐injury dosing paradigm congruent with its mechanisms of action, proved efficacious in reversing functional impairment after TBI in developing rats.

Support or Funding Information

Dedicated health research funds from the University of New Mexico, Genise Goldenson Foundation and Boston Children's Hospital.
Oxytocin Protects Against Oxidative Stress And Apoptosis‐Induced Cell Damage In Glial “U87‐MG” Cell‐Line: Role Of OXT Receptor
Authors: ALANAZI MOHAMMED , Havranek Tomas , Bakos Jan , Castejon Ana Maria
Published: November 2023

Background

the neuropeptide oxytocin, released from the posterior pituitary into the systemic circulation has been implicated in several vital physiological processes, ranging from reproduction to social and non‐social behaviors. Oxytocin has been associated with cell proliferation, protection against apoptosis, and regulation of cytoskeletal protein expression. These findings might be relevant since certain neurological disorders have been linked to low systemic levels of oxytocin (e.g. autism).

Objective

we evaluated both the anti oxidant capacity and the protective role of oxytocin against apoptosis and oxidative stress in the Glioblastoma “U87‐MG” cell line.

Methodology

we developed an Oxytocin Receptor Knockdown cell line, U87MG_KD OXT‐R (astrocyte), to further analyze 1) viability rates after different stressors (H2O2, Camptothecin or DMNQ) with and without oxytocin for 24 hours 2) Anti oxidant capacity of oxytocin against H2O2 for 24 hours.

Results

In the control cell‐line “U87‐MG”, treatment with 100nM and 1uM oxytocin significantly blunted the reduction in cell viability induced by H2O2 (p<0.01, p<0.001), Camptothecin (p<0.05, p<0.01) and DMNQ (p<0.05, p<0.001) respectively. Also, oxytocin (100 nM and 1uM) significantly reduced reactive oxygen species ROS generation when cells were exposed to 500uM H2O2 (p<0.05 and p<0.05 respectively). In contrast, oxytocin did not protect against cell loss or affected ROS production in U87MG_KD OXT‐R when cells were treated with these oxidative stress and pro‐apoptotic substances.

Conclusion

our results indicate that oxytocin has a protective and anti‐oxidative effect in U87‐MG and this effect is lacking when the OXTR is knockdown. These results contribute to a better understanding of oxytocin's effect and its potential role in many neurological disorders associated with abnormal cell viability and oxidative stress such as autism.

Support or Funding Information

1‐ President's Faculty Research & Development (PFRDG) Grant ‐ 2015–2016

2‐ Saudi Arabian Cultural Mission (SACM) Graduate Students Research Grant