Vehicle Solution Research Articles

Therapeutic effects of cinnamon bark oil on sciatic nerve injury in rats.

The aim of this study was to investigate the effects of cinnamon bark essential oil (CBO) on analgesia, motor activity, balance, and coordination in rats with sciatic nerve damage. Rats were divided into three groups as simply randomized. The right sciatic nerve (RSN) of the Sham group was explored. Only vehicle solution was applied for 28 days. The RSN of the sciatic nerve injury (SNI) group was explored. Damage was created by unilateral clamping, and vehicle solution was applied for 28 days. The RSN of the sciatic nerve injury+cinnamon bark essential oil (SNI+CBO) group was explored. SNI was created by unilateral clamping and CBO was applied for 28 days. In the experiment study, motor activity, balance, and coordination measurements were made with rotarod and accelerod tests. A hot plate test was performed for analgesia measurements. Histopathology studies were carried out with the sciatic nerve tissues. In the rotarod test, there was a statistically significant difference between the SNI group and the SNI+CBO group (p<0.05). According to the accelerod test findings, there was a statistically significant difference between the SNI group with the Sham and SNI+CBO groups. In the hot plate test, there was a statistically significant difference between the SNI group with the Sham and SNI+CBO groups (p<0.05). In comparison to the Sham group and the SNI group, the SNI+CBO group was shown to have the greatest expression level of vimentin. We have concluded that CBO can be used as an adjuvant treatment in cases of SNI, increased pain, nociception, impaired balance, motor activity, and coordination. Our results will be supported by further studies.

Therapeutic effect of C-type natriuretic peptide on persistent pain in a rat knee arthritis model.

Intra-articular injection of C-type natriuretic peptide (CNP) at the acute inflammatory stage suppressed fibrotic changes in the infrapatellar fat pad (IFP), articular cartilage degeneration, and persistent pain in a monoiodoacetic acid (MIA)-induced rat knee arthritis model. In this study, we administered CNP during the inflammation subsiding period to evaluate CNP effectiveness in knees with osteoarthritis (OA) pathology. 20male Wistar rats were randomly divided into two groups. The rats received an intra-articular injection of MIA solution in the right knee to induce inflammation-induced joint degeneration. One group subsequently received an intra-articular CNP injection for six consecutive days from day 8, whereas another group received vehicle solution. Pain avoidance behavior tests and histological analyses were conducted to examine the therapeutic effects of CNP. The incapacitance test indicated that the percent weight on the ipsilateral limb decreased after MIA injection by day 4 and continued to decrease until the end of the experiment in the vehicle group, suggesting persistent pain in the knee. Intra-articular injection of CNP reversed the weight-bearing ratio on day 19. Histological evaluation showed that the CNP group had more residual fat tissue in the IFP and fewer calcitonin gene-related peptide-positive nerve endings compared to the vehicle group. CNP could not reverse articular cartilage degeneration. Intra-articular injection of CNP after the IFP fibrosis onset had no significant effect on OA severity and extent. Nevertheless, CNP might be utilized therapeutically for OA treatment since it can alleviate persistent knee pain and inhibit structural changes in residual fat tissue.

Non-psychoactive Cannabidiol Prevents Osteoporosis in an Animal Model and Increases Cell Viability, Proliferation, and Osteogenic Gene Expression in Human Skeletal Stem and Progenitor Cells.

Cannabidiol (CBD), the non-psychoactive component of the Cannabis sativa plant, is marketed as a potential therapeutic agent and has been studied for its roles in reducing inflammation and managing neuropathic pain. Some studies have reported that CB1 and CB2 receptor activation can attenuate and reverse bone loss in experimental animal models. Despite this, little is known about the impact of CBD on fracture healing. We investigated the effects of CBD in vitro using human osteoprogenitor cells and in vivo via murine femur fracture and osteoporosis models. In vitro mesenchymal stem cells were treated with increasing concentrations of crystalized pharmaceutical grade CBD or vehicle solution. Cell viability and proliferation were significantly increased in cells treated with CBD compared to vehicle control. Osteocalcin expression was also significantly higher in the CBD-treated human stem cells compared to vehicle control. In vivo the effect of CBD on bone mineral density and fracture healing in mice was examined using a two-phase experimental approach. Fluoxetine was used for pharmacologic induction of osteoporosis and surgical oophorectomy (OVX) was used for hormonal induction of osteoporosis. X-ray and microCT analysis showed that CBD prevented both fluoxetine- and OVX-induced osteoporosis. We found that while OVX resulted in delayed bone healing in control mice, CBD-pretreated mice exhibited normal bone healing. Collectively these in vitro and in vivo findings suggest that CBD exerts cell-specific effects which can be exploited to enhance bone metabolism. These findings also indicate that CBD usage in an osteoporotic population may positively impact bone morphology, warranting further research.

Adaptive Optics Flood Illumination Ophthalmoscopy in Nonhuman Primates: Findings in Normal and Short-term Induced Detached Retinae

To describe adaptive optics flood illumination ophthalmoscopy (AO-FIO) of the photoreceptor layer in normal nonhuman primates (NHPs) and in the case of a short-term induced retinal detachment (RD). Longitudinal fundamental research study. Four NHPs were used to image normal retinae with AO-FIO (in comparison with 4 healthy humans); 2 NHPs were used to assess the effects of RD. The photoreceptor layer (cone mosaic metrics, including cone density, cone spacing, and cone regularity) was followed with AO-FIO imaging (rtx1, Imagine Eyes) during a surgically induced RD in 2 NHPs using a vehicle solution containing dimethyl sulfoxide, classically used as a chemical solvent. We also performed functional testing of the retina (full-field and multifocal electroretinogram [ERG]). Correlation of cone mosaic metrics (cone density, spacing, and regularity) between normal retinae of NHPs and humans, and cone metrics, power spectrum, and ERG wave amplitudes after RD. Imaging features were very similar in terms of cone reflectivity, cell density, regularity, and spacing values, showing strong positive correlations between NHPs and humans. After RD, AO-FIO revealed several alterations of the cone mosaic slowly recovering during the 3 months after the reattachment, which were not detected functionally by ERG. These results demonstrate by invivo AO-FIO imaging the transient structural changes of photoreceptors after an RD in the primate retina. They also provide an interesting illustration of the AO-FIO potential for investigating photoreceptor toxicity during preclinical studies in NHPs with a high translatability to human studies. Proprietary or commercial disclosure may be found after the references.

Acute effects of empagliflozin on open-loop baroreflex function and urine glucose excretion in Goto-Kakizaki diabetic rats

Although suppression of sympathetic activity is suggested as one of the underlying mechanisms for the cardioprotective effects afforded by sodium–glucose cotransporter 2 (SGLT2) inhibitors, whether the modulation of glucose handling acutely affects sympathetic regulation of arterial pressure remains to be elucidated. In Goto–Kakizaki diabetic rats, we estimated the open-loop static characteristics of the carotid sinus baroreflex together with urine glucose excretion using repeated 11-min step input sequences. After the completion of the 2nd sequence, an SGLT2 inhibitor empagliflozin (10 mg kg−1) or vehicle solution was administered intravenously (n = 7 rats each). Empagliflozin did not significantly affect the baroreflex neural or peripheral arc, despite significantly increasing urine glucose excretion (from 0.365 ± 0.216 to 8.514 ± 0.864 mg·min−1·kg−1, P < 0.001) in the 7th and 8th sequences. The possible sympathoinhibitory effect of empagliflozin may be an indirect effect associated with chronic improvements in renal energy status and general disease conditions.

Role of the iNOS isoform in the cardiovascular dysfunctions of male rats with 6-OHDA-induced Parkinsonism

IntroductionAvailable studies have shown the involvement of nitric oxide (NO) in the processes that lead to neurodegeneration in Parkinson's disease (PD). Also, the use of inhibitors of the inducible isoform of NO-synthase (iNOS) promotes neuroprotection and attenuates dopamine (DA) loss in experimental models of Parkinsonism. In addition, NO also appears to be involved in cardiovascular changes in 6-hydroxydopamine (6-OHDA)-induced Parkinsonism. The current study aimed to evaluate the effects of iNOS inhibition on cardiovascular and autonomic function in animals that were subjected to Parkinsonism by the administration of 6-OHDA. Materials and methodsThe animals underwent stereotaxic surgery for bilateral microinfusion of the neurotoxin 6-OHDA (6 mg/mL in 0.2% ascorbic acid in sterile saline solution) or vehicle solution for the Sham group. From the day of stereotaxis until the day of femoral artery catheterization, the animals were treated with the iNOS inhibitor, S-methylisothiourea (SMT; 10 mg/kg; i. p.) or saline solution (0.9%; i. p.) for 7 days. The animals were divided into four groups: Sham-Saline, Sham-SMT, 6-OHDA-Saline, and 6-OHDA-SMT. Subsequent analyses were performed on these four groups. After 6 days, they underwent catheterization of the femoral artery, and 24 h later, mean arterial pressure (MAP) and heart rate (HR) were recorded. Another group of animals (the 6-OHDA and Sham groups) was assessed for aortic vascular reactivity after 7 days of bilateral infusion of 6-OHDA or vehicle, in which cumulative concentration-effect curves (CCEC) were made for phenylephrine (Phenyl), acetylcholine and sodium nitroprusside (NPS). Also, CCEC in the presence of Nw-nitro-arginine-methyl-ester (l-NAME) (10-5 M), SMT (10-6 M), and indomethacin (10-5 M) blockers were made. ResultsThe effectiveness of the 6-OHDA lesion was confirmed with the reduction of DA in 6-OHDA animals. However, treatment with SMT could not reverse the loss of DA. Concerning the baseline parameters, SBP and MAP values were lower in 6-OHDA animals compared to their Sham control, with no effect of treatment with SMT. In the analysis of SBP variability, a decrease in variance, the VLFabs component, and the LFabs component were observed in the 6-OHDA groups when compared to their controls, regardless of treatment with SMT. It was also observed that intravenous injections of SMT resulted in an increase in BP and a decrease in HR. However, the response was not different between the Sham and 6-OHDA groups. In vascular function, there was a hyporeactivity to Phenyl in the 6-OHDA group, and when investigating the mechanisms of this hyporeactivity, it was seen that the Rmax to Phenyl increased with incubation with SMT, indicating that iNOS could be involved in the vascular hyporeactivity of animals with Parkinsonism. ConclusionThus, the set of results presented in this study suggests that part of the cardiovascular dysfunction in animals subjected to 6-OHDA Parkinsonism may be peripheral and involve the participation of endothelial iNOS.

Effects of Resveratrol on Vascular Function in Retinal Ischemia-Reperfusion Injury.

Ischemia-reperfusion (I/R) events are involved in the development of various ocular pathologies, e.g., retinal artery or vein occlusion. We tested the hypothesis that resveratrol is protective against I/R injury in the murine retina. Intraocular pressure (IOP) was elevated in anaesthetized mice to 110 mm Hg for 45 min via a micropipette placed in the anterior chamber to induce ocular ischemia. In the fellow eye, which served as control, IOP was kept at a physiological level. One group received resveratrol (30 mg/kg/day p.o. once daily) starting one day before the I/R event, whereas the other group of mice received vehicle solution only. On day eight after the I/R event, mice were sacrificed and retinal wholemounts were prepared and immuno-stained using a Brn3a antibody to quantify retinal ganglion cells. Reactivity of retinal arterioles was measured in retinal vascular preparations using video microscopy. Reactive oxygen species (ROS) and nitrogen species (RNS) were quantified in ocular cryosections by dihydroethidium and anti-3-nitrotyrosine staining, respectively. Moreover, hypoxic, redox and nitric oxide synthase gene expression was quantified in retinal explants by PCR. I/R significantly diminished retinal ganglion cell number in vehicle-treated mice. Conversely, only a negligible reduction in retinal ganglion cell number was observed in resveratrol-treated mice following I/R. Endothelial function and autoregulation were markedly reduced, which was accompanied by increased ROS and RNS in retinal blood vessels of vehicle-exposed mice following I/R, whereas resveratrol preserved vascular endothelial function and autoregulation and blunted ROS and RNS formation. Moreover, resveratrol reduced I/R-induced mRNA expression for the prooxidant enzyme, nicotinamide adenine dinucleotide phosphate oxidase 2 (NOX2). Our data provide evidence that resveratrol protects from I/R-induced retinal ganglion cell loss and endothelial dysfunction in the murine retina by reducing nitro-oxidative stress possibly via suppression of NOX2 upregulation.

Collagen matricryptin promotes cardiac function by mediating scar formation

AimsA peptide mimetic of a collagen-derived matricryptin (p1159) was shown to reduce left ventricular (LV) dilation and fibrosis after 7 days delivery in a mouse model of myocardial infarction (MI). This suggested p1159 long-term treatment post-MI could have beneficial effects and reduce/prevent adverse LV remodeling. This study aimed to test the potential of p1159 to reduce adverse cardiac remodeling in a chronic MI model and to elucidate p1159 mode-of-action. Materials and methodsUsing a permanent occlusion MI rodent model, animals received p1159 or vehicle solution up to 28 days. We assessed peptide treatment effects on scar composition and structure and on systolic function. To assess peptide effects on scar vascularization, a cohort of mice were injected with Griffonia simplicifolia isolectin-B4. To investigate p1159 mode-of-action, LV fibroblasts from naïve animals were treated with increasing doses of p1159. Key findingsMatricryptin p1159 significantly improved systolic function post-MI (2-fold greater EF compared to controls) by reducing left ventricular dilation and inducing the formation of a compliant and organized infarct scar, which promoted LV contractility and preserved the structural integrity of the heart. Specifically, infarcted scars from p1159-treated animals displayed collagen fibers aligned parallel to the epicardium, to resist circumferential stretching, with reduced levels of cross-linking, and improved tissue perfusion. In addition, we found that p1159 increases cardiac fibroblast migration by activating RhoA pathways via the membrane receptor integrin α4. SignificanceOur data indicate p1159 treatment reduced adverse LV remodeling post-MI by modulating the deposition, arrangement, and perfusion of the fibrotic scar.

Early DNase-I therapy delays secondary brain damage after traumatic brain injury in adult mice

Traumatic brain injury (TBI) causes the release of danger-associated molecular patterns (DAMP) from damaged or dead cells, which contribute to secondary brain damage after TBI. Cell-free DNA (cfDNA) is a DAMP known to cause disruption of the blood–brain barrier (BBB), promote procoagulant processes, brain edema, and neuroinflammation. This study tested the hypothesis that administration of deoxyribonuclease-I (DNase-I) has a beneficial effect after TBI. Mice (n = 84) were subjected to controlled cortical impact (CCI) and posttraumatic intraperitoneal injections of low dose (LD) or high dose (HD) of DNase-I or vehicle solution at 30 min and 12 h after CCI. LD was most effective to reduce lesion volume (p = 0.003), brain water content (p < 0.0001) and to stabilize BBB integrity (p = 0.019) 1 day post-injury (dpi). At 6 h post injury LD-treated animals showed less cleavage of fibrin (p = 0.0014), and enhanced perfusion as assessed by micro-computer-tomography (p = 0.027). At 5 dpi the number of Iba1-positive cells (p = 0.037) were reduced, but the number of CD45-positive cells, motoric function and brain lesion volume was not different. Posttraumatic-treatment with DNase-I therefore stabilizes the BBB, reduces the formation of brain edema, immune response, and delays secondary brain damage. DNase-I might be a new approach to extend the treatment window after TBI.

The Vagus Nerve Mediates Gut-Brain Response to Duodenal Nutrient Administration.

Obesity contributes significant disease burden worldwide, including diabetes, cardiovascular disease, and cancer. While bariatric surgery is the most effective and durable obesity treatment, the mechanisms underlying its effects remain unknown. Although neuro-hormonal mechanisms have been suspected to mediate at least some of the gut-brain axis changes following bariatric surgery, studies examining the intestine and its regionally specific post-gastric alterations to these signals remain unclear. Vagus nerve recording was performed following the implantation of duodenal feeding tubes in mice. Testing conditions and measurements were made under anesthesia during baseline, nutrient or vehicle solution delivery, and post-delivery. Solutions tested included water, glucose, glucose with an inhibitor of glucose absorption (phlorizin), and a hydrolyzed protein solution. Vagus nerve signaling was detectable from the duodenum and exhibited stable baseline activity without responding to osmotic pressure gradients. Duodenal-delivered glucose and protein robustly increased vagus nerve signaling, but increased signaling was abolished during the co-administration of glucose and phlorizin. Gut-brain communication via the vagus nerve emanating from the duodenum is nutrient sensitive and easily measurable in mice. Examination of these signaling pathways may help elucidate how the nutrient signals from the intestine are altered when applied to obesity and bariatric surgery mouse models. Future studies will address quantifying the changes in neuroendocrine nutrient signals in health and obesity, with specific emphasis on identifying the changes associated with bariatric surgery and other gastrointestinal surgery.

How to achieve sustainable distribution in the fast fashion industry? An electric vehicle solution under the "vehicle-battery separation" mode.

Exploring sustainable urban distribution based on electric vehicles is crucial given the rise in global greenhouse gas emissions, especially for fast fashion industries with extremely high distribution frequencies. However, most studies have overlooked the impact of deep discharge on the distribution route scheme, and few studies fit the characteristics of the fast fashion industry. As a result, this study presents a novel electric vehicle routing problem considering deep discharge model (EVRP-DD) for distribution route optimization, which fully considers deep discharge under the emerging mode of vehicle-battery separation. The characteristics of fashion consumers and products were also integrated into the model, such as consumer satisfaction and 3D loading constraints. To solve this complex programming problem, a sophisticated hybrid ant colony optimization (HACO) algorithm was designed by combining the advantages of ACO and A-star algorithms. Using real-life data, the experimental results verify the effectiveness and superiority of the proposed solution. EVRP-DD achieved reduced driving distance, total distribution cost, and deep discharge distance. HACO can enhance the computation speed and reduce the total distribution cost compared with the two conventional algorithms. The proposed solution showed excellent flexibility and could effectively adjust the optimal route scheme according to the ever-changing external environment. Thus, it can be concluded that this solution is a powerful tool for enterprises to achieve sustainable distribution. This study has realized theoretical innovation in sustainable distribution under the new mode of vehicle-battery separation, and its successful application in the fast fashion industry reflects its valuable application value.

Effects of C1-INH Treatment on Neurobehavioral Sequelae and Late Seizures After Traumatic Brain Injury in a Mouse Model of Controlled Cortical Impact.

C1 human-derived C1 esterase inhibitor (C1-INH) is a U.S. Food and Drig Administration-approved drug with anti-inflammatory actions. In the present study, we investigated the therapeutic effects of C1-INH on acute and chronic neurobehavioral outcomes and on seizures in the chronic stage in a mouse traumatic brain injury (TBI) model. Adult male CD1 mice were subjected to controlled cortical impact and randomly allocated to receive C1-INH or vehicle solution 1 h post-TBI. Effects of C1-INH treatment on inflammatory responses and brain damage after TBI were examined using the Cytometric Bead Array, C5a enzyme-linked immunosorbent assay, Fluoro-Jade C staining, and Nissl staining. Neurobehavioral outcomes after TBI were assessed with modified neurological severity scores, the rotarod and open field tests, and the active place avoidance task. Video-electroencephalographic monitoring was performed in the 15th and 16th weeks after TBI to document epileptic seizures. We found that C1-INH treatment reduced TNFα expression and alleviated brain damage. Treatment with C1-INH improved neurological functions, increased locomotor activity, alleviated anxiety-like behavior, and exhibited an effect on seizures in the chronic stage after TBI. These findings suggest that C1-INH has beneficial effects on the treatment of TBI.

Effect of Liposomal-Lactoferrin-Based Eye Drops on the Conjunctival Microflora of Patients Undergoing Cataract Surgery.

Postoperative endophthalmitis is typically caused by the patient's conjunctival bacterial flora. Povidone iodine solution (5%) is used perioperatively to obtain periocular and ocular antisepsis. However, an adjunctive prophylaxis procedure could further help control the conjunctival microbial load. Considering the increase in antibiotic resistance, a progressive shift toward alternative methods would be desirable. Somilux® eye drops (Alfa Intes, lactoferrin-based eye drops) are medical devices containing liposomal lactoferrin (LF). This study evaluates the effects on conjunctival microflora of LF-based eye drops used in the preoperative phase in patients scheduled for cataract surgery. LF-based eye drops or a vehicle solution (water solution) were instilled 4 times a day starting 3days before cataract surgery. Before the therapy (T0) and at the time of surgery (T1), a conjunctival swab was performed in both eyes and processed to detect microbial growth, microbiological isolation, and species identification. The outcome was the quantification and characterization of the local microbial flora before and after using LF-based or vehicle-based eye drops. Safety of the treatments was also evaluated. 88 eyes of 44 patients (mean [± SD] age 75 [± 12.6] years) were enrolled. At baseline, 54 conjunctival swabs showed only saprophytic flora, 27 showed only potential pathogenic flora, and seven showed both of them. LF-based eye drops reduced the proportion of potentially pathogenic bacteria (36% at T0 vs. 9% at T1, p = 0.008) compared with the vehicle (41% at T0 vs. 55% at T1, p = 0.302) without altering the physiological ocular microbial composition. No adverse events have been reported. Our findings provide a novel contribution to the scientific knowledge on the role of LF in the ophthalmic field, supporting the use of LF-based eye drops as a safe and selective treatment to improve the ocular surface physiological defenses and control the bacterial ocular surface contamination prior to cataract surgery.

Gastrodin ameliorates atherosclerosis by inhibiting foam cells formation and inflammation through down-regulating NF-κB pathway

BackgroundGastrodin is an effective polyphenol extracted from Chinese natural herbal Gastrodiae elata Blume, which exhibits antioxidant and anti-inflammatory effects. It has been reported to benefit neurodegenerative diseases, but the effect of Gastrodin on atherosclerosis and the underlying mechanisms remain elusive. The aim of this study is to investigate the function and mechanism of Gastrodin in atherosclerosis.MethodsAtherosclerosis mouse model was established by fed low density lipoprotein receptor-deficient (Ldlr−/−) mice with a high fat diet (HFD, 20% fat and 0.5 cholesterol) for 8 weeks and Gastrodin was administered daily via oral gavage. Plasma lipid levels were measured using commercial kits. En face and aortic sinus lipid accumulation were analyzed with Oil Red O staining. In vitro cell models using foam cell formation model and classical atherosclerosis inflammation model, macrophages were incubated with oxygenized low-density lipoproteins (ox-LDL) or lipopolysaccharide (LPS) in the presence of different concentration of Gastrodin or vehicle solution. Foam cell formation and cellular lipid content were evaluated by Oil Red O staining and intracellular lipids extraction analysis. Gene expression and proteins related to cholesterol influx and efflux were examined by quantitative reverse transcription PCR (RT-qPCR) and western blotting analysis. Furthermore, the effect of Gastrodin on LPS induced macrophage inflammatory responses and NF-κB pathway were evaluated by RT-qPCR and western blotting analysis.ResultsGastrodin administration reduced the body weight, plasma lipid levels in Ldlr−/− mice after fed a high fat diet. Oil Red O staining showed Gastrodin-treated mice displayed less atherosclerosis lesion area. Furthermore, Gastrodin treatment significantly ameliorated ox-LDL-induced macrophage-derived foam cells formation through suppressing genes expression related to cholesterol efflux including scavenger receptor class B and ATP-binding cassette transporter A1. Moreover, Gastrodin markedly suppressed pro-inflammatory cytokines secretion and LPS induced inflammatory response in macrophage through downregulating NF-κB pathway.ConclusionsOur study demonstrated that Gastrodin attenuates atherosclerosis by suppressing foam cells formation and LPS-induced inflammatory response and represents a novel therapeutic target for the treatment of atherosclerosis.

Transient depletion of macrophages alters local inflammatory response at the site of disc herniation in a transgenic mouse model

Macrophages are abundantly detected at sites of disc herniation, however, their function in the disease progression is unclear. We aim to investigate the functions of macrophages in acute disc herniation using a macrophage Fas-induced apoptosis (MaFIA) transgenic mouse strain. To transiently deplete macrophages, a dimerizer, AP20187, or vehicle solution was administered via intraperitoneal injection to MaFIA mice immediately, day 1 and 2 after annular puncture induced disc herniation. Local infiltrated tissues at disc hernia and DRGs at corresponding levels were harvested to analyze immune cells and neuroinflammation on postoperative day (POD) 6 by flow cytometry and/or immunostaining. Mouse spines were harvested to analyze structures of degenerated discs and adjacent vertebrae and to assess osteoclast activity by histology and tartrate-resistant acid phosphatase (TRAP) staining on POD 6, 13, and 20, respectively. On POD 6, abundant macrophages were confirmed at disc hernia sites. Compared to vehicle control, AP20187 significantly reduced GFP+ cells in blood, spleen, and local inflammatory tissue. At disc hernia sites, AP20187 markedly reduced macrophages (CD11b+, F4/80+, GFP+CD11b+, CD11b+F4/80+) while increasing neutrophils and B cells. Transient macrophage depletion decreased ectopic bone formation and osteoclast activity in herniated discs and adjacent cortical bones for up to 20 days post herniation. Disc herniation elevated expressions of TNF-α, IL-6, substance P, calcitonin gene-related peptide, accompanied by increasing GFP+, CD11b+ and F4/80+ macrophages. Macrophage depletion did not attenuate these markers of neuroinflammation. Transient depletion of macrophages altered local inflammatory response at the site of disc herniation.

Physicochemical stability of durvalumab (Imfinzi®) concentrate for solution in original vials after first opening

Abstract Objectives Durvalumab (Imfinzi®), a PD-L1 monoclonal antibody (mAb) medication is available as concentrate (50 mg/mL) for solution for infusion. The summary of product characteristics provides information about the physicochemical stability of ready-to-administer durvalumab preparations (vehicle solution 0.9 % NaCl, G5%), but not about the concentrate after first opening. The objective of this study was to determine the physicochemical stability of durvalumab concentrate for solution after first opening over a period of 28 days. Methods Imfinzi® vials were punctured and stored refrigerated (2–8 °C) or at room temperature (20–25 °C) light protected. At predefined time points (day 0, 7, 14, 21, 28) the physicochemical stability of the concentrated solution was determined by ion-exchange/size-exclusion high-performance liquid chromatography (IE-/SE-HPLC) with photodiode array detection and pH measurement. Vials were inspected with regard to changes of color, clarity, and visible particles at any time point. Results Regardless of the storage temperature, durvalumab 50 mg/mL solutions remained physiochemically stable for 28 days in punctured vials. The concentrations of durvalumab monomer remained unchanged and no secondary peaks (fragments, aggregates) were observed in any of the SE-HPLC chromatograms. The IE-HPLC test results showed no substantial changes of the peak areas of the main peak and of the acidic and basic charge variants during the whole storage period. Appearance and pH of the test solutions remained unchanged until the end of the study. Conclusions Regardless of storage conditions none of the analytical methods indicated physicochemical instability of the intact durvalumab monomer over the 28 days of the study. To avoid microbiological instability storage under refrigeration is recommended.

Physicochemical Stability of Generic Thiotepa Concentrate and Ready-to-Administer Infusion Solutions for Conditioning Treatment

The objective of this study was to determine the physicochemical in-use stability of recently approved Thiotepa Riemser concentrate in the original vial and diluted ready-to-administer (RTA) infusion solutions in prefilled glucose 5% and 0.9% NaCl polyolefin bags. Thiotepa Riemser 10 mg/mL concentrates and infusion solutions (1 mg/mL, 2 mg/mL, 3 mg/mL) were prepared in triplicate and stored at 2-8 °C or 25 °C for 14 days. Thiotepa concentrations were determined using a stability-indicating RP-HPLC assay. In parallel, pH and osmolality were measured. Sub-visible particles were counted on day 0 and 14. Thiotepa Riemser concentrate was revealed to be stable for 14 days when stored at 2-8 °C, or for 24 h when stored at 25 °C. Thiotepa concentrations in infusion solutions stored at 2-8 °C remained above 95% of the initial concentrations for at least 14 days, regardless of the type of vehicle solution. When stored at 25 °C, thiotepa infusion solutions in glucose 5% proved to be physicochemically stable for 3 days (1 mg/mL), 5 days (2 mg/mL) or 7 days (3 mg/mL). Thiotepa infusion solutions in 0.9% NaCl remained physicochemically stable for 5 days (1 mg/mL) or 7 days (2 mg/mL, 3 mg/mL). At these points in time, the specification limit of ≤0.6% monochloro-adduct was fulfilled. In parallel, an elevation of the pH values was registered. Thiotepa concentrates and infusion solutions should be stored at 2-8 °C due to temperature-dependent physicochemical stability, and for microbiological reasons. Glucose 5% infusion solution is recommended as a diluent, and stability-improving nominal 2 mg/mL to 3 mg/mL thiotepa concentrations should be obtained.

Role of synthetical amynoquinone ethyl 2-(1,4-dioxo-1,4-dihydronaphthalen-2-ylamino) acetate in inhibition of Ehrlich’s tumor

Quinones are naturally or synthetically occurring secondary metabolites that have various bio-dynamics, highlighting their antitumor potential. This has been explored through their selective cytotoxicity, and studies in medicinal chemistry about the relation between biological activity versus chemical structure may lead to the solution of the toxicity problems associated with quinones. In this context, the antitumor effect of a synthetic naphthoquinone, named Ethyl 2-(1,4-Dioxo-1,4-Dihydronaphthalen-2-Ylamino) Acetate, was tested using mice transplanted with Ehrlich ascitic tumor as an experimental model. The acute toxicity test was performed using 30 mice that received the aminoquinone at doses of 100, 200, 300, and 600mg/kg. After evaluation of the clinical findings in the spontaneous activity tests, the LD50 calculation for the test substance showed low levels of toxicity at doses lower than 244.11±23.29mg/kg. Thus, three experimental groups were established, where animals transplanted with tumor cells received NaCl vehicle solution (control, n=6), and the others were treated with 71.7mg/kg of Methotrexate (n=6) or 20mg/kg of Aminoquinone (n=6). All administrations were intraperitoneal, in a single dose. Three days after the implantation of the tumor cells the animals were weighed daily and evaluated for tumor biometry and development. The treatments occurred five days after the implantation of the tumor cells and were extended for 7 more days. At the end of the 12-day experimental period, all animals were euthanized for biochemical and histopathological analyses of the tumors and vital organs. The spontaneous activity test showed that the amount of responses associated with the nervous system tends to increase with the increase in dosage, highlighting the excitatory effect on the central nervous system in almost all dosages employed, followed by depressant activities on this system. There was a significant tumor reduction, both in animals treated with methotrexate (71.7 %) and in those treated with aminoquinone (91.6 %) in the control group. There was no significant difference in tumor volume between the animals treated with aminoquinone or methotrexate. The histopathological analysis revealed that in both treatments there were fewer mitoses in the tumor mass compared to the control group. However, there was apparent toxicity to the liver, heart, and left kidney in the treatment with methotrexate compared to aminoquinone. The significant capacity for tumor reduction presented by aminoquinone allows pointing it as a promising alternative for the development of a more efficient drug to control tumor development, being necessary for the development of new studies to deepen the knowledge about its mechanisms of action.

The Role of Intra-Amygdaloid Neurotensin and Dopamine Interaction in Spatial Learning and Memory.

Neurotransmitter and neuromodulator neurotensin (NT) has been proved to facilitate spatial and passive avoidance learning after microinjected into the rat central nucleus of amygdala (CeA). These previous studies of our laboratory also revealed that neurotensin-1 receptor (NTS1) is involved in the mentioned actions of NT. Extensive literature confirms the interaction between neurotensinergic and dopaminergic systems, and our research group also suppose that the mesolimbic dopaminergic system (MLDS) is involved in the spatial learning and memory-facilitating effect of NT in the CeA. In the present work, NT and dopamine (DA) interaction has been examined in the Morris water maze and passive avoidance tests. Rats received 100 ng NT, 5 µg dopamine D2 receptor antagonist sulpiride in itself, sulpiride as a pretreatment before NT or vehicle solution into the CeA. NT microinjection significantly decreased target-finding latency in the Morris water maze test and significantly increased entrance latency in the passive avoidance test, as was expected based on our previous findings. The DA D2 receptor antagonist pretreatment was able to inhibit both effects of NT. The results confirm the facilitatory effect of NT on spatial learning and memory and let us conclude that these actions can be exerted via the DA D2 receptors.

Osmoprotective and bioprotective effect of threalose 3% under desiccating conditions: An in vitro study

AbstractPurpose: To study the osmoprotective and bioprotective effect of trehalose 3% during desiccation conditions.Methods: Normal human conjunctival epithelial cells (IOBA‐NHC) were exposed to: 1) culture media DMEM‐F12 (Control); 2) Carboxymethylcellulose 0.5% + glycerin 0.9% + PEG 400 0.25% (Vehicle); 3) Vehicle + trehalose 3% (Trehalose). Cultures were treated for 1 h and then exposed to desiccating conditions (25°C and 36–38% relative humidity). Metabolic activity by MTT assay and cell viability by Triptan Blue assay was evaluated. To evaluate water retention capacity, a gravimetric test was carried out by soaking a filter paper with 5 ml of each solution under evaluation and recording its weight over time. The data were analysed by analysis of variance (ANOVA) with a post hoc Tukey multiple comparisons test.Results: MTT assay: During the first 30 min under desiccating conditions there were no differences between groups (Control: 0.586 ± 0.012; Vehicle: 0.575 ± 0.278; Trehalose: 0.523 ± 0.035). After 45 min of exposure metabolic activity sharply decreased in control group (0.111 ± 0.007) and vehicle group (0.107 ± 0.027), while trehalose group maintained metabolic activity values (0.560 ± 0.008; p &lt; 0.001).Triptan Blue assay: The cell viability of the trehalose group was maintained throughout the time (10 min: 62.7 ± 2.5; 15 min: 58.7 ± 8.1; 20 min: 62.3 ± 10.8; 30 min:41.7 ± 8.5%) and was statistically superior to the control and vehicle groups at all evaluated times (p &lt; 0.01).Water retention capacity: Water content of papers soaked with control and vehicle solution was almost completely evaporated (less than 20% of water retention) at 10 min on desiccating conditions, while trehalose group show a significant delay of water evaporation of 5 and 7 min regarding the vehicle and control group, respectively.Conclusions: The addition of trehalose 3% prolonged cell viability and extended water retention time in conditions of high environmental dryness. This study confirms the bioprotective and osmoprotective role of trehalose evaluated in previous investigations and provides scientific evidence of its properties, representing an additional benefit in the treatment of dry eye.