Injection Of Phosphate-buffered Saline Research Articles

Evaluation of experimental animal behaviors through establishment of an ovalbumin-induced experimental mouse model of allergic rhinitis

Allergic rhinitis (AR) was also called hay fever which was a type of nasal inflammation when the immune system overreacts to environmental allergen exposures. AR’s clinical symptoms included a runny or stuffy nose, sneezing, red, itchy, watery eyes, and eye swelling. The fluid in the nasal cavity was usually clear. Patients with AR can affect sleep and work qualities. Seriously, the AR symptoms can also cause asthma, allergic conjunctivitis, or atopic dermatitis. Therefore, it is an important issue to attenuate AR symptoms and research the novel therapeutic drugs for AR patients. The purpose of this study was to introduce an easy-to-establish experimental mouse model of AR. In this study, the male BALB/c mice were divided respectively into as the Group A (n = 12) and the Group B (n = 12). Group A and Group B were designed as the normal control and RA, respectively. BALB/c mice in Group B were sensitized by intraperitoneal injection of ovalbumin (OVA) on day 0, day 4, day 13, and day 20, followed by continuous nasal administration of OVA solution once per day between day 21-43. BALB/c mice in Group A received sensitization of intraperitoneal injection of phosphate-buffered saline (PBS) on day 0, day 4, day 13, and day 20 and continuous nasal administration of PBS instead of OVA once per day between day 21-43. Before and after sensitization, the frequencies of nasal symptoms (sneezing, nasal rubbing) were recorded and counted. Results were showed that sneezing times in Group B were higher than Group A on D29, D30, D36, and D43 of the experiment. The sneezing times in Group A were significant higher on D29 and D30 of the experiment. However, the sneezing times in Group B were significant higher on D29, D30, D36, and D43 of the experiment. The rubbing times in Group B were higher than Group A on D29, D30, D36, and D43 of the experiment. The rubbing times in Group A were significant higher on D30 and D43 of the experiment. However, the rubbing times in Group B were significant higher on D29, D30, D36, and D43 of the experiment. Based on these results, a successful mouse model of AR has been established. We hope that this RA mouse model will provide a tool for the research of the novel AR therapeutic drugs and apply these novel AR therapeutic drugs to attenuate the AR symptoms in AR patients in the future.

A fat body transcriptome analysis of the immune responses of Rhodnius prolixus to artificial infections with bacteria

BackgroundRhodnius prolixus is an important vector of Trypanosoma cruzi, the causal agent of Chagas disease in humans. Despite the medical importance of this and other triatomine vectors, the study of their immune responses has been limited to a few molecular pathways and processes. Insect immunity studies were first described for holometabolous insects such as Drosophila melanogaster, and it was assumed that their immune responses were conserved in all insects. However, study of the immune responses of triatomines and other hemimetabolous insects has revealed discrepancies between these and the Drosophila model.MethodsTo expand our understanding of innate immune responses of triatomines to pathogens, we injected fifth instar nymphs of R. prolixus with the Gram-negative (Gr−) bacterium Enterobacter cloacae, the Gram-positive (Gr+) bacterium Staphylococcus aureus, or phosphate-buffered saline (PBS), and evaluated transcript expression in the fat body 8 and 24 h post-injection (hpi). We analyzed the differential expression of transcripts at each time point, and across time, for each treatment.ResultsAt 8 hpi, the Gr− bacteria-injected group had a large number of differentially expressed (DE) transcripts, and most of the changes in transcript expression were maintained at 24 hpi. In the Gr+ bacteria treatment, few DE transcripts were detected at 8 hpi, but a large number of transcripts were DE at 24 hpi. Unexpectedly, the PBS control also had a large number of DE transcripts at 24 hpi. Very few DE transcripts were common to the different treatments and time points, indicating a high specificity of the immune responses of R. prolixus to different pathogens. Antimicrobial peptides known to be induced by the immune deficiency pathway were induced upon Gr− bacterial infection. Many transcripts of genes from the Toll pathway that are thought to participate in responses to Gr+ bacteria and fungi were induced by both bacteria and PBS treatment. Pathogen recognition receptors and serine protease cascade transcripts were also overexpressed after Gr− bacteria and PBS injections. Gr- injection also upregulated transcripts involved in the metabolism of tyrosine, a major substrate involved in the melanotic encapsulation response to pathogens.ConclusionsThese results reveal time-dependent pathogen-specific regulation of immune responses in triatomines, and hint at strong interactions between the immune deficiency and Toll pathways.Graphical abstract

Quantitative cerebrovascular reactivity MRI in mice using acetazolamide challenge.

To develop a quantitative MRI method to estimate cerebrovascular reactivity (CVR) in mice. We described an MRI procedure to measure cerebral vasodilatory response to acetazolamide (ACZ), a vasoactive agent previously used in human clinical imaging. Vascular response was determined by cerebral blood flow (CBF) measured with phase-contrast or pseudo-continuous arterial spin labeling MRI. Vasodilatory input intensity was determined by plasma ACZ level using high-performance liquid chromatography. We verified the source of the CVR MRI signal by comparing ACZ injection to phosphate-buffered saline injection and noninjection experiments. Dose dependence and feasibility of regional CVR measurement were also investigated. Cerebral blood flow revealed an exponential increase following intravenous ACZ injection, with a time constant of 1.62 min. In contrast, phosphate-buffered saline or noninjection exhibited a slow linear CBF increase, consistent with a gradual accumulation of anesthetic agent, isoflurane, used in this study. When comparing different ACZ doses, injections of 30, 60, 120, and 180 mg/kg yielded a linear increase in plasma ACZ concentration (p < 0.0001). On the other hand, CBF changes under these doses were not different from each other (p=0.50). The pseudo-continuous arterial spin labeling MRI with multiple postlabeling delays revealed similar vascular responses at different postlabeling delay values. There was a regional difference in CVR (p=0.005), with isocortex (0.81 ± 0.17%/[μg/ml]) showing higher CVR than deep-brain regions. Mice receiving multiple ACZ injections lived for a minimum of 6months after the study without noticeable aberrant behavior or appearance. We demonstrated the proof-of-principle of a new quantitative CVR mapping technique in mice.

A Single Injection of NTG-101 Reduces the Expression of Pain-Related Neurotrophins in a Canine Model of Degenerative Disc Disease

Background: Tissue sources of pain emanating from degenerative discs remains incompletely understood. Canine intervertebral discs (IVDs) were needle puncture injured, 4-weeks later injected with either phosphate-buffered saline (PBS) or NTG-101, harvested after an additional fourteen weeks and then histologically evaluated for the expression of NGFr, BDNF, TrkB and CALCRL proteins. Quantification was performed using the HALO automated cell-counting scoring platform. Immunohistochemical analysis was also performed on human IVD tissue samples obtained from spinal surgery. Immunohistochemical analysis and quantification of neurotrophins and neuropeptides was performed using an in vivo canine model of degenerative disc disease and human degenerative disc tissue sections. Discs injected with NTG-101 showed significantly lower levels of Nerve Growth Factor receptor (NGFr/TrkA, p = 0.0001), BDNF (p = 0.009), TrkB (p = 0.002) and CALCRL (p = 0.008) relative to PBS injections. Human IVD tissue obtained from spinal surgery due to painful DDD show robust expression of NGFr, BDNF, TrkB and CALCRL proteins. A single intradiscal injection of NTG-101 significantly inhibits the expression of NGFr, BDNF, TrkB and CALCRL proteins in degenerative canine IVDs. These results strongly suggest that NTG-101 inhibits the development of neurotrophins that are strongly associated with painful degenerative disc disease and may have profound effects upon the management of patients living with discogenic pain.

The Preparation, Biodistribution, and Dosimetry of Encapsulated Radio-Scandium in a Dendrimer for Radio-nano-pharmaceutical Application.

This study aimed to investigate the synthesis, characterization, and biodistribution of scandium nanoparticles encapsulated within poly (amidoamine) (PAMAM) dendrimers, as well as to estimate the human absorbed dose. It also aimed to examine, in particular, the amine-terminated PAMAM dendrimers in generation 5. Irradiation of the compound in the nuclear reactor resulted in the formation of Sc-radioactive complex nanoparticles. The compound of the dendrimer-Sc3+ was confirmed by the UV-vis spectrometer. The size of the particles was less than 10 nm, and it was assessed using high-resolution transmission electron microscopy (HRTEM) and dynamic light scattering (DLS). The synthesized complex was irradiated by the 3 × 1011 n.cm-2s-1 flux of neutron for 2 h. Mice bearing a breast tumor were employed to assess the therapeutic dose that was delivered by the poly scandium-46-nanoparticles. As opposed to the untreated groups, a single injection of poly phosphate-buffered saline to intratumoral in other groups to deliver a dose of 100 µCi resulted in a statistically significant 39.24% reduction in tumor volume 14 days after injection. After applying the biokinetics data in mice, the human’s absorbed dose from scandium-47 encapsulated PAMAM was extrapolated based on animal data. The absorbed doses in critical organs, including the liver, lung, spleen, kidney, and bone, were 0.879, 0.0472, 0.191, 0.107, and 0.155 mGy/MBq, respectively.

Electromyographic characterization and bioinformatic pathway analysis of spinal cord inflammation in a Tat induced HIV-associated sensory neuropathy model

Abstract HIV sensory neuropathy (HIV-SN) is the most prevalent neurological complication of HIV infection. The contribution of Tat in HIV-SN has not been studied extensively. Previously, doxycycline (DOX) inducible HIV-1 Tat transgenic (iTat) mice have been shown to exhibit neuropathy-like behaviors following Tat induction. To further understand Tat-associated HIV-SN, here, we first characterized the model via electromyographic (EMG) evaluation. EMG testing of the hind leg along the sciatic nerve was conducted before DOX (daily i.p. for 14 days at 100 mg/kg) or vehicle (pH-matched phosphate-buffered saline) injection and at days 3, 7, 14, 21, 28 and 35 post the first injection. Initial analysis showed a slightly increased maximal amplitude in DOX mice along with the decreased hind paw grip strength. Further, we performed a NanoString RNA assessment of lumbar spinal cord tissues using a panel of 200+ inflammation-related genes, which was followed by a series of bioinformatic pathway analyses. Short Time-Series Expression Miner (STEM) cluster analysis identified four significant temporal expression patterns of genes. These four profiles were characterized by continuous downregulation, early-, mid-, and late-upregulation. Using PANTHER GO-pathway analysis we found that three signaling pathways were common amongst the four STEM profiles which were the apoptosis signaling, inflammation mediated by chemokine and cytokine signaling, and the Toll-like receptor signaling pathways. Previous studies highlighted selected genes that belong to these pathways and are also targets of Tat. Future work will focus on investigating how these pathways contribute to sensory neuropathy and identifying biomarkers for HIV-SN. Supported by NIH/NIDA 1R21DA044886 (Cao)

Possible interaction of central noradrenergic, serotoninergic and oxytocin systems with nesfatin-1 induced hypophagia and feeding behavior in newborn broiler

There are some differences between mammals and birds in terms of central food intake regulation. In avian species, the hypophagic role of nesfatin-1 has not been investigated with other neurotransmitters. Therefore, this study aimed to determine the alteration of feeding behavior following intracerebroventricular (ICV) injection of nesfatin-1 and its possible interaction with central noradrenergic, serotoninergic, and oxytocin systems in newborn broiler chicks. In experiment 1, birds received ICV injection of phosphate-buffered saline (PBS), prazosin (α1 receptors antagonist, 10 nmol), nesfatin-1 (40 ng), and co-administration of prazosin and nesfatin-1. Experiments 2–10 were similar to experiment 1, except that yohimbine (α2 receptors antagonist, 13 nmol), metoprolol (β1 receptors antagonist, 24 nmol), IC1118,551 (β2 receptors antagonist for, 5nmol), SR59230R (β3 receptors antagonist, 20 nmol), fluoxetine (serotonin reuptake inhibitor, 10 µg), PCPA (serotonin synthesis inhibitor, 1.5 µg), 8-OH-DPAT (5-HT1A receptors agonist, 15.25 nmol), SB242084 (5-HT2C receptors antagonist,1.5 µg) and tocinoic acid (oxytocin receptors antagonist, 2 µg) were injected instead of prazosin. Immediately after the injection, food consumption and behavioral traits were recorded. Nesfatin-1 decreased food consumption (P < 0.05). Nesfatin-1 along with ICI118551 decreased food consumption (P < 0.05). The nesfatin-1- induced hypophagia were reduced by the simultaneous injection of PCPA and nesfatin-1 (P < 0.05). Nesfatin-1induced hypophagia were decreased by the simultaneous injection of SB242084 (P < 0.05). The nesfatin-1 -induced hypophagia were abolished by the simultaneous injection of the tocinoic acid and nesfatin-1 (P < 0.05). ICV injection of the nesfatin-1 decreased the number of steps, jumps, exploratory food, and pecks (P < 0.05) with no effect on drink pecks (P > 0.05). Nesfatin-1 significantly decreased standing time and increased both sitting time and rest time (P < 0.05). Nesfatin-1 could play an important role in feeding behavior, and its hypophagic effects were mediated by β2 adrenergic, 5-HT2C serotoninergic, and oxytocin receptors in neonatal chickens.

Anti-CD80/86 antibodies inhibit inflammatory reaction and improve graft survival in a high-risk murine corneal transplantation rejection model

We investigated the effects of anti-CD80/86 antibodies in a murine high-risk corneal transplantation rejection model. A mixed lymphocyte reaction (MLR) assay was conducted with anti-CD80/86 antibodies. Inflammatory cytokine levels in the culture supernatant were measured using an enzyme-linked immunosorbent assay. Interferon (IFN)-γ-producing CD4+ T cell frequencies in the MLR were assessed using flow cytometry. In vivo, high-risk corneal allograft survival and IFN-γ-producing CD4+ T cell frequencies in corneal grafts were assessed with intraperitoneal injection of anti-CD80/86 antibodies compared to phosphate-buffered saline (PBS). RNA-sequencing was performed on corneal grafts 2 weeks post-transplantation. Anti-CD80/86 antibodies significantly decreased T-cell proliferation, IFN-γ+-producing CD4+ T cell frequencies, and IFN-γ, interleukin (IL)-1β, IL-2, IL-10, and tumor necrosis factor-α production in the MLR compared to PBS injection. Intraperitoneal injection of anti-CD80/86 antibodies significantly prolonged corneal graft survival and decreased IFN-γ+-producing CD4+ T cell frequencies compared to PBS injection. Gene set enrichment analysis showed that the gene sets mainly enriched in the control group were related to allograft rejection and inflammatory response compared to PBS injection. Anti-CD80/86 antibodies significantly prolonged corneal graft survival by inhibiting T-cell proliferation and inflammatory response.

The Learning Curve of Murine Subretinal Injection Among Clinically Trained Ophthalmic Surgeons.

PurposeSubretinal injection (SRI) in mice is widely used in retinal research, yet the learning curve (LC) of this surgically challenging technique is unknown.MethodsTo evaluate the LC for SRI in a murine model, we analyzed training data from three clinically trained ophthalmic surgeons from 2018 to 2020. Successful SRI was defined as either the absence of retinal pigment epithelium (RPE) degeneration after phosphate buffered saline injection or the presence of RPE degeneration after Alu RNA injection. Multivariable survival-time regression models were used to evaluate the association between surgeon experience and success rate, with adjustment for injection agents, and to calculate an approximate case number to achieve a 95% success rate. Cumulative sum (CUSUM) analyses were performed and plotted individually to monitor each surgeon's simultaneous performance.ResultsDespite prior microsurgery experience, the combined average success rate of the first 50 cases in mice was only 27%. The predicted SRI success rate did not reach a plateau above 95% until approximately 364 prior cases. Using the 364 training cases as a cutoff point, the predicted probability of success for cases 1 to 364 was 65.38%, and for cases 365 to 455 it was 99.32% (P < 0.0001). CUSUM analysis showed an initial upward slope and then remained within the decision intervals with an acceptable success rate set at 95% in the late stage.ConclusionsThis study demonstrates the complexity and substantial LC for successful SRI in mice with high confidence. A systematic training system could improve the reliability and reproducibility of SRI-related experiments and improve the interpretation of experimental results using this technique.Translational RelevanceOur prediction model and monitor system allow objective quantification of technical proficiency in the field of subretinal drug delivery and gene therapy for the first time, to the best of our knowledge.

Association between anti-fibrillin-2 protein induced retinal degeneration via intravitreous delivery and activated TGF-β signalling in mice.

Fibrillin-2 (FBN2) is a major component of tissue microfibrils, and the decrease of FBN2 perturbs the signalling events mediated by transforming growth factor-β (TGF-β), thereby playing a role in macular degeneration. However, the association between the retinal degeneration resulting from the abnormality of FBN2 and the activation of TGF-β signalling has not been fully addressed. In the present study, the mice were divided into a normal control group (NC group), a phosphate-buffered saline (PBS) injection group (PBS group), and an anti-FBN2 protein injection group (anti-FBN2group), and the mice in PBS and anti-FBN2groups received the relevant treatment via the intravitreal injection once a week for three consecutive weeks. One week later after injection, the retinal morphology and visual function of the fundus were detected. Further, the expression of FBN2, TGF-β1, TGF-β2 and TGF-β3 in retina was measured using quantitative polymerase chain reaction and enzyme-linked immunosorbent assay (ELISA), respectively. As a result, fundus examination suggests that after intravitreous injection of anti-FBN2 protein, there were a large patchy yellow white degeneration region and numerous pigmentations in the retina in anti-FBN2-treated mice; by contrast, there was no apparent change in mice from the NC and PBS groups. The retina suffered markedly damage, and the thickness of whole retina and outer nuclear layer markedly thinned. The expression of FBN2 was decreased whereas the levels of TGF-β1, TGF-β2 and TGF-β3 were upregulated. Together, our findings indicate that the intravitreous delivery of anti-FBN2 protein could induce retina degeneration in mice, accompanied by the higher activated TGF-β. The retinal degeneration mouse model established will provide a platform for the investigation of the retinal diseases.

Hedgehog-Gli2 Signaling Promotes Chemoresistance in Ovarian Cancer Cells by Regulating MDR1.

BackgroundCisplatin (DDP) resistance remains a key challenge in improving the clinical outcome of patients with ovarian cancer (OC). Gli2 overexpression can lead to DDP resistance in OC cells, but the specific underlying regulatory mechanism remains unclear. The membrane transporter encoding gene MDR1 positively regulates chemotherapy resistance in various cancer types. We evaluated MDR1 as a potential Gli2 downstream target and the contribution of the Gli2/MDR1 axis in promoting DDP resistance in OC cells.MethodsTo generate drug-resistant SKOV3/DDP cells, SKOV3 cells were grown for six months under continuous induction wherein the DDP concentration was steadily increased. Gli2 expression in OC cells with varying DDP sensitivities was detected using western blot. Cell counting kit-8 assays were used to assess the DDP sensitivity of SKOV3, SKOV3/DDP, A2780, and A2780/DDP cells and reversal of DDP resistance in SKOV3/DDP and A2780/DDP cells. Cell proliferation was analyzed using 5-ethynyl-2′-deoxyuridine (EdU) incorporation assays. The transcriptional regulation of MDR1 by Gli2 was determined using luciferase reporter assays. Finally, xenograft OC tumors were generated in nude mice, which were then treated with intraperitoneal DDP or phosphate-buffered saline (PBS) injections to investigate if Gli2 affected DDP resistance in OC in vivo.ResultsDDP-resistant SKOV3/DDP and A2780/DDP cells showed higher expression of Gli2 and MDR1 as compared with that in DDP-sensitive OC cells. Gli2 knockdown in SKOV3/DDP cells significantly reduced MDR1 expression, whereas it increased DNA damage, thereby sensitizing OC cells to DDP. Similar results were obtained after targeting Gli2 expression with the Gli-antagonist 61 inhibitor (GANT61) in SKOV3/DDP and A2780/DDP cells. In cells stably overexpressing Gli2, treatment with gradient concentrations of verapamil, an MDR1 inhibitor, significantly inhibited MDR1 expression. Our findings indicate that downregulation of MDR1 expression may reverse OC cell resistance to DDP. Moreover, dual-luciferase reporter gene assays confirmed that MDR1 is a direct downstream target of Gli2, with Gli2 positively regulating MDR1 expression. Finally, subcutaneous xenotransplantation in nude mice demonstrated that Gli2 plays a key role in regulating OC drug resistance.ConclusionsWe identified a mechanism by which Hedgehog-Gli signaling regulates OC chemoresistance by modulating MDR1 expression. Hence, Gli2 and MDR1 are potential biomarkers and therapeutic targets in patients with chemoresistant OC.

Prenatal infection predisposes offspring to enhanced susceptibility to imiquimod-mediated psoriasiform dermatitis in mice

Background: Infection can upregulate T helper 17 (Th17) signaling and exacerbate psoriasis. Literature has indicated that prenatal infection induces embryonic development toward Th17 signaling and is a risk factor for developing certain Th17-skewing disorders such as autism in offspring. However, it remains unclear if a prenatal infection is a risk factor for developing psoriasis in offspring. Objectives: We investigated if a prenatal infection predisposes mice offspring to enhanced susceptibility to psoriatic inflammation. Methods: Pregnant C57BL/6 mice were intraperitoneally injected with lipopolysaccharide (LPS) or polyinosinic:polycytidylic acid poly I:C to simulate prenatal bacterial and viral infection, respectively. Phosphate-buffered saline (PBS) injection was used as the control. When mice offspring were at the age of 9–10 weeks, imiquimod (IMQ) cream or control vehicle was applied topically on ears for 5 consecutive days to induce psoriasiform dermatitis. The extent of epidermal hyperplasia and the number of Munro microabscesses were examined by histologic analysis. Protein expressions of neutrophil marker Ly6 g and Th17-associated cytokines were measured by western blotting. Results: Prenatal infection with LPS or poly I:C induced a greater extent of epidermal hyperplasia and more Munro microabscesses after IMQ application as compared with prenatal PBS injection. Mice offspring with prenatal infection also had higher protein expressions of Ly6 g, IL-17a, and interleukin-23 after IMQ stimulation compared with their PBS controls. Conclusion: Prenatal infection predisposes mice offspring to enhanced susceptibility to IMQ-mediated psoriasiform dermatitis. The data obtained from the present animal study suggest that prenatal infection might be a risk factor for developing psoriasis in offspring.

Pericyte-derived heme-binding protein 1 promotes angiogenesis and improves erectile function in diabetic mice

PurposeTo comprehensively evaluate the effect on angiogenesis of heme-binding protein 1 (Hebp1) in the treatment of diabetes-induced erectile dysfunction.Materials and MethodsMouse corpus cavernosum endothelial cells and pericytes were used for in vitro study. Four groups of mice were used: control nondiabetic mice and streptozotocin-induced diabetic mice receiving two intracavernous injections of phosphate-buffered saline, Hebp1 (1 µg), or Hebp1 (5 µg). The function of Hebp1 in diabetic conditions was evaluated by tube formation assay, aorta ring assay, migration assay, intracavernous pressure, immunofluorescence staining, and Western blot experiments.ResultsWe report that Hebp1 is more highly expressed in mouse corpus cavernosum pericytes and can effectively promote endothelial cell angiogenesis under high-glucose conditions. Following exogenous administration of Hebp1 protein, we found that elevated Hebp1 levels can improve the erectile function of diabetic mice, which is achieved by reducing reactive oxygen species levels and activating the PI3K/AKT/eNOS signaling pathway.ConclusionsOur findings demonstrate that Hebp1 can promote angiogenesis and improve erectile function under diabetic conditions.

Insulin gene expression and functional activity of insulin signaling pathway in Alzheimer's disease

Aim. To study the insulin (INS) gene expression, insulin and lactate levels, expression of Fe65 adapter protein, and level of oxidative DNA damage marker γH2AX in different brain areas in the experimental model of Alzheimer's disease.Materials and Methods. Male, 4-month-old C57BL/6 mice received either intrahippocampal injection of β-amyloid (C57BL/6 + Aβ 1-42) or phosphate-buffered saline (C57BL/6 + PBS). Insulin (INS) gene expression in the hippocampus and amygdala was assessed by means of reverse transcription-polymerase chain reaction. Levels of lactate and insulin in different brain areas were measured by enzyme-linked immunosorbent assay. Expression of Fe65 adapter protein and γH2AX in the hippocampus was studied by immunofluorescence staining followed by confocal microscopy.Results. We found an overexpression of the INS gene in the hippocampus and amygdala, an increase in lactate level in the hippocampus, and slightly increased insulin level in the amygdala of mice with Alzheimer's disease as compared with the control group. Neurodegeneration was accompanied by an elevated endothelial expression of Fe65 adapter protein (p= 0.04) and γH2AX in hippocampal neurons (p = 0.04).Conclusion. Alzheimer's disease neurodegeneration is accompanied by a disrupted insulin signaling and impaired glucose metabolism in the hippocampus and amygdala. This further leads to a neuronal accumulation of γH2AX and impaired amyloid precursor protein proteolysis because of insulin inability to inhibit its interaction with the Fe65 adapter protein and to prevent formation and deposition of β-amyloid.

Application of Hyaluronic Acid as a Biopolymer Material in Reconstruction of Interdental Papilla in Rats

Applying hyaluronic acid, a biopolymer material, in the treatment of interdental papilla reconstruction has become a trend. The main objective of this research is to investigate the histologic effect of hyaluronic acid on interdental papilla over time. Deficient interdental papilla models were surgically created in sixty-two Sprague-Dawley (SD) rats and were randomly treated with the injection of hyaluronic acid (HA group) or phosphate-buffered saline (sham control group) or left untreated (control group). After 2, 4, and 8 weeks, the rats were sacrificed in batches to observe the histological changes. A fluorochrome label was used to monitor bone formation in 8 weeks. Immunohistochemical analyses were performed to evaluate the expression of potentially relevant cytokines, vascular endothelial growth factor (VEGF), bone morphogenetic protein-2 (BMP-2), and Wnt-induced secreted protein 1 (WISP1) in the gingival tissue in 8 weeks. A preliminary study of HA degradation after 24 weeks was performed in two rats. Following the HA injection, no inflammation or granulomatous foreign body reaction was observed. HA was able to promote collagen fiber and alveolar bone regular formation in the reconstruction site. HA also enhanced VEGF, BMP-2, and WISP-1 expression in gingival tissue (p＜0.05). After 24 weeks, there was no HA filler observed in the interdental papilla. In conclusion, our study suggested that HA is an effective way to reconstruct deficient interdental papilla.

Effects of remote ischemic preconditioning on the deformability and aggregation of red blood cells in a rat endotoxemia model.

The prevention of rheologic alterations in erythrocytes may be important for reducing sepsis-associated morbidity and mortality. Remote ischemic preconditioning (RIPC) has been shown to prevent tissue damage caused by severe ischemia and mortality resulting from sepsis. However, the effect of RIPC on erythrocytes in sepsis is yet to be determined. To investigate the effect of RIPC on rheologic alterations in erythrocytes in sepsis. Thirty male Sprague-Dawley rats were used in this study. An endotoxin-induced sepsis model was established by intraperitoneally injecting 20 mg/kg LPS (LPS group). RIPC was induced in the right hind limb using a tourniquet, with three 10-minute of ischemia and 10 min of reperfusion cycles immediately before the injection of LPS (RIPC/LPS group) or phosphate-buffered saline (RIPC group). The aggregation index (AI), time to half-maximal aggregation (T1/2), and maximal elongation index (EImax) of the erythrocytes were measured 8 h after injection. The AI, T1/2, and EImax values in the LPS and RIPC/LPS groups differed significantly from those in the RIPC group, but there were no differences between the values in the LPS and RIPC/LPS groups. RIPC did not prevent rheologic alterations in erythrocytes in the rat model of LPS-induced endotoxemia.

Анализ ретинальной экспрессии генов, участвующих в регуляции ангиогенеза и функций эндотелиального барьера после воздействия лазерного излучения с длиной волны 577 нм в непрерывном режиме на сетчатку

Objective. The aim of the study was to investigate the retinal expression of genes involved in the regulation of angiogenesis and endothelial barrier function after exposure of the retina to 577 nm laser light in a continuous mode in experimental conditions after intravitreal injection of VEGF165 (Vascular endothelial growth factor). Materials and methods. The study was performed on 4-5 week old male C57BL / 6J mice. 4 groups, 5 mice in each, were formed, one animal eye was experimental and the second remained intact. The animals of the first group received intravitreal injection of phosphate-buffered saline (PBS); animals in the second, third and fourth groups received intravitreal injection of 50 ng/ml of recombinant VEGF165; in the third and fourth groups, one day after the VEGF165 intravitreal administration, laser irradiation with a wavelength of 577 nm was performed on the retina in micropulse and continuous modes, respectively. Tissue samples from animals of the first and second groups were taken 2 days after PBS and VEGF165 administration, and in animals from the third and fourth groups one day after laser exposure to the retina. Results. Retinal expression of genes involved in the regulation of angiogenesis and endothelial barrier function as a result of retinal exposure to 577 nm laser radiation in continuous mode following intravitreal injection of VEGF165 was studied. Genes with significant changes in expression levels were identified (genes that regulate the processes of angiogenesis). Conclusion. Understanding the mechanisms of modulation of retinal gene expression may help to identify the key regulatory factors providing therapeutic effects of laser radiation in continuous and micropulsed modes and provide the basis for future therapeutic tactics for retinal diseases.

Human Adipose-Derived Mesenchymal Stem Cells Ameliorate Elastase-Induced Emphysema in Mice by Mesenchymal-Epithelial Transition.

PurposeChronic obstructive pulmonary disease (COPD) is a worldwide problem because of its high prevalence and mortality. However, there is no fundamental treatment to ameliorate their pathological change in COPD lung. Recently, adipose-derived mesenchymal stem cells (ADSCs) have attracted attention in the field of regenerative medicine to repair damaged organs. Moreover, their utility in treating respiratory diseases has been reported in some animal models. However, the detailed mechanism by which ADSCs improve chronic respiratory diseases, including COPD, remains to be elucidated. We examined whether human ADSCs (hADSCs) ameliorated elastase-induced emphysema and whether hADSCs differentiated into alveolar epithelial cells in a murine model of COPD.MethodsFemale SCID-beige mice (6 weeks old) were divided into the following four groups according to whether they received an intratracheal injection of phosphate-buffered saline or porcine pancreatic elastase, and whether they received an intravenous injection of saline or hADSCs 3 days after intratracheal injection; Control group, hADSC group, Elastase group, and Elastase-hADSC group. We evaluated the lung function, assessed histological changes, and compared gene expression between hADSCs isolated from the lung of Elastase-hADSC group and naïve hADSCs 28 days after saline or elastase administration.ResultshADSCs improved the pathogenesis of COPD, including the mean linear intercept and forced expiratory volume, in an elastase-induced emphysema model in mice. Furthermore, hADSCs were observed in the lungs of elastase-treated mice at 25 days after administration. These cells expressed genes related to mesenchymal–epithelial transition and surface markers of alveolar epithelial cells, such as TTF-1, β-catenin, and E-cadherin.ConclusionhADSCs have the potential to improve the pathogenesis of COPD by differentiating into alveolar epithelial cells by mesenchymal–epithelial transition.

Amniotic fluid stem cell administration can prevent epithelial injury from necrotizing enterocolitis.

Stem cell therapy has been proven to rescue intestinal injury and stimulate intestinal regeneration in necrotizing enterocolitis (NEC). Specifically, stem cells derived from amniotic fluid (AFSCs) and mesenchymal stem cells (MSCs) derived from bone marrow have shown promising results in the treatment of experimental NEC. This study aims to examine the effects of AFSCs and MSCs on the prevention of intestinal injury during experimental NEC. Supernatants from AFSC and MSC cultures were collected to perform proteomic analysis. Prior to NEC induction, mice received intraperitoneal injections of phosphate-buffered saline (PBS), 2 × 106 AFSCs, or 2 × 106 MSCs. We found that AFSCs grew faster than MSCs. Proteomic analysis indicated that AFSCs are primarily involved in cell development and growth, while MSCs are involved in immune regulation. Administering AFSCs before NEC induction decreased NEC severity and mucosal inflammation. Intestinal proliferation and endogenous stem cell activation were increased after AFSC administration. However, administering MSCs before NEC induction had no beneficial effects. This study demonstrated that AFSCs and MSCs have different protein release profiles. AFSCs can potentially be used as a preventative strategy for neonates at risk of NEC, while MSCs cannot be used. AFSCs and MSCs have distinct protein secretory profiles, and AFSCs are primarily involved in cell development and growth, while MSCs are involved in immune regulation. AFSCs are unique in transiently enhancing healthy intestinal epithelial cell growth, which offers protection against the development of experimental NEC. The prevention of NEC via the administration of AFSCs should be evaluated in infants at great risk of developing NEC or in infants with early signs of NEC.

Effect and Mechanism of Treating Experimental Autoimmune Encephalomyelitis in Mice with Butylphthalide Combined with Bone Marrow Mesenchymal Stem Cells

To explore the efficacy and mechanism of using 3-n-butylphthalide (NBP) in combination with bone marrow mesenchymal stem cells (BMSCs) in the treatment of experimental autoimmune encephalomyelitis (EAE) in mice. Myelin oligodendrocyte glycoprotein (MOG35-55) was used for the induction and establishment of the EAE model in C57BL/6 mice. The mice were randomly assigned to the EAE group, which received intraperitoneal injection of phosphate-buffered saline (PBS), the NBP-treated EAE group, or the NBP group, which received intraperitoneal injection of NBP, the BMSCs transplantion EAE group, or the BMSCs group, which received BMSCs injected into the lateral ventricle and intraperitoneal injection of PBS, and the BMSCs and NBP combination treatment EAE group, or the BMSCs+NBP group, which received BMSCs injected into the lateral ventricle and intraperitoneal injection of NBP. Each group had 10 mice, while ten normal mice were used as the blank control group receiving intraperitoneal injection of PBS. The neurological function scores were documented daily. The mice were sacrificed 22 days after EAE induction, and the demyelination state of of the spinal cords was observed through Luxol fast blue (LFB) staining. In addition, the levels of serum interleukin-6 (IL-6), IL-10, IL-17, IL-22 and transforming growth factor-β (TGF-β) were examined with ELISA. The levels of glial fibrillary acidic protein (GFAP), microtubule associated protein-2 (MAP-2) and myelin basic protein (MBP) in the brain were examined with immunofluorescence staining. Western blot was used to check the expressions of nuclear factor (NF)-κB pathway, phosphoinositide-3 kinase (PI3K)/protein kinase B (PKB or Akt) pathway, IL-17 and forkhead box P3 (Foxp3) in the spinal cords. The neurological function scores and average scores of each treatment group were significantly lower than those of the EAE group ( P<0.05). The scores of the BMSCs+NBP group decreased more significantly than those of the single treatment groups (the NBP group and the BMSCs group) ( P<0.05). LFB staining results of the spinal cords were consistent with the neurological function scores and the average scores. Compared with the EAE group, the levels of pro-inflammatory cytokines, including IL-6, IL-17 and IL-22, significantly decreased ( P<0.05), and the levels of anti-inflammatory cytokines IL-10 and TGF-β significantly increased ( P<0.05). The change in cytokine expression was more significant in the BMSCs+NBP group ( P<0.05). The expressions of GFAP, MAP-2 and MBP in the BMSCs+NBP group were significantly higher than those of the BMSCs group ( P<0.05). Compared with the EAE group, the p-NF-κB/NF-κB ratio and the IL-17/Foxp3 ratio in NBP group, BMSCs group and BMSCs+NBP group decreased, while P-IκBα/IκBα, p-pI3k/PI3K and P-Akt/Akt ratios increased, especially in the BMSCs+NBP group( P<0.05). The combined treatment of NBP and BMSCs can help alleviate the symptoms of EAE model mice, showing better efficacy than treatment with NBP or BMSCs alone. The mechanism is related to the inhibition of the NF-κB pathway to regulate Th17/Foxp3 ratio and the activation of the PI3K/Akt pathway to promote the neurogenic differentiation of BMSCs.