Journal: Brain Sciences

Article Title: Caloric Restriction Mimetic 2-Deoxyglucose Reduces Inflammatory Signaling in Human Astrocytes: Implications for Therapeutic Strategies Targeting Neurodegenerative Diseases

doi: 10.3390/brainsci12030308

Figure Lengend Snippet: 2-DG blocks the expression of multiple IL-1β-induced inflammatory genes in human astrocytes. Fold change of IL-1β ( A ), TNFα ( B ), C3 ( C ), or LCN2 ( D ) mRNA transcript levels normalized to ACTB mRNA levels in total RNA isolated from human astrocytes. One-way ANOVA was performed to the determine effect of treatment on IL-1β [F (9, 10) = 505.5, p < 0.0001], TNFα [F (9, 10) = 96.72, p < 0.0001], C3 [F (9, 10) = 796.1, p < 0.0001], and LCN2 [F (9, 10) = 516.8, p < 0.0001]. A post hoc Tukey’s test was conducted with corrected p -values shown (** p < 0.01; *** p < 0.001 vs. vehicle; ^^^ p < 0.001 vs. IL-1β-treated cells).

Article Snippet: Gene expression was determined using RT 2 PCR TaqMan gene expression assays with the QuantStudio 3 sequence-detections system (Life Technologies TM ) using Taqman primers specific to IL-1β (cat. no. Hs01555410_m1), IL6 (cat. no. Hs00174131_m1), TNFα (cat. no. Hs00174128_m1), C3 (cat. no. Hs00163811_m1), and LCN2 (cat. no. Hs01008571_m1).

Techniques: Expressing, Isolation

Journal: Brain Sciences

Article Title: Caloric Restriction Mimetic 2-Deoxyglucose Reduces Inflammatory Signaling in Human Astrocytes: Implications for Therapeutic Strategies Targeting Neurodegenerative Diseases

doi: 10.3390/brainsci12030308

Figure Lengend Snippet: 2-DG blocks IL-1β-induced inflammatory gene expression in human astrocytes. IL-1β is a prototypic inflammatory cytokine that is relevant to many neurodegenerative diseases. IL-1β was used here to model neuroinflammation while 2-DG was used to mimic caloric restriction, a phenomenon relevant to fasting and exercise, both of which may have beneficial effects on the brain. Our findings indicate that 2-DG reduces the IL-1β-induced expression of inflammatory genes in astrocytes including, but not limited to, IL6, TNFα, C3, and LCN2. This finding may partially explain the beneficial effects of exercise, caloric restriction, and ketogenic diets. Future studies will further investigate the mechanisms through which 2-DG is reducing astrocyte inflammatory gene expression as well as the downstream effects on neuronal functioning and blood–brain barrier integrity.

Article Snippet: Gene expression was determined using RT 2 PCR TaqMan gene expression assays with the QuantStudio 3 sequence-detections system (Life Technologies TM ) using Taqman primers specific to IL-1β (cat. no. Hs01555410_m1), IL6 (cat. no. Hs00174131_m1), TNFα (cat. no. Hs00174128_m1), C3 (cat. no. Hs00163811_m1), and LCN2 (cat. no. Hs01008571_m1).

Techniques: Gene Expression, Expressing

Journal: Brain

Article Title: Connexin-43 induces chemokine release from spinal cord astrocytes to maintain late-phase neuropathic pain in mice

doi: 10.1093/brain/awu140

Figure Lengend Snippet: Spinal injection of TNF-α-activated astrocytes induces mechanical allodynia via Cx43-mediated CXCL1 release. (A) Intrathecal injection of TNF-α-activated astrocytes elicited persistent mechanical allodynia for >48 h. Note this allodynia is reduced by pretreatment of astrocytes with Cx43 small interfering RNA (1 µg/ml, 18 h). *P < 0.05, compared with TNF-α or TNF-α + non-targeting control small interfering RNA treated group; n = 6 mice/group. (B) ELISA analysis shows increased CXCL1 release in the CSF at 3 h after the intrathecal injection of TNF-α-activated astrocytes. *P < 0.05, compared with vehcile group; #P < 0.05, compared with non-activated astrocytes; n = 4 mice/group. (C) Intrathecal injection of a CXCL1 neutralizing antibody (4 µg) transiently and partially reversed mechanical allodynia, induced by TNF-α-treated astrocytes. *P < 0.05, compared with control IgG group; n = 6 mice/group. (D) Intrathecal injection of the CXCR2 antagonist SB225002 (20 µg = 57 nmol) transiently and partially reversed mechanical allodynia, induced by TNF-α-activated astrocytes. *P < 0.05, compared with vehicle (PBS); n = 5–6 mice/group. All data are mean ± SEM. The differences between groups were analysed by ANOVA followed by Newman–Keuls test.

Article Snippet: The sections were then incubated overnight at 4°C with the following primary antibodies: GFAP antibody (1:1000, mouse; Millipore Bioscience Research Reagents), Cx43 antibody (1:1000, rabbit; Sigma), NeuN antibody (1:1000, mouse; Millipore Bioscience Research Reagents), CXCL1 (1: 200, rabbit; Boster), and CXCR2 antibody (1: 200, rabbit; Boster).

Techniques: Injection, Small Interfering RNA, Control, Enzyme-linked Immunosorbent Assay

Journal: Brain

Article Title: Connexin-43 induces chemokine release from spinal cord astrocytes to maintain late-phase neuropathic pain in mice

doi: 10.1093/brain/awu140

Figure Lengend Snippet: CXCL1, upregulated in spinal cord astrocytes after nerve injury, enhances excitatory synaptic transmission in spinal cord neurons and maintains neuropathic pain via CXCR2. (A) Western blotting shows CXCL1 upregulation in the spinal cord dorsal horn 21 days after CCI. Right, quantification of Cx43 levels in the dorsal horn. The western blot results are presented as a fold of sham control. *P < 0.05, compared to sham control, Student’s t-test, n = 4 mice/group. (B) Intrathecal injection of SB 225002 (20 µg), 21 days after CCI, reduced CCI-induced mechanical allodynia in the late phase. *P < 0.05, compared with vehicle (saline), Student’s t-test, n = 6 mice/group. (C) Double immunostaining of CXCL1 and GFAP in the dorsal horn 21 days after CCI. Note CXCL1 is primarily colocalized with GFAP. Arrows indicate doubled-labelled cells. Scale bar = 50 µm. (D and E) CXCL1 superfusion (100 ng/ml) increases spontaneous EPSC frequency (revealed by patch clamp recordings) in lamina IIo neurons of spinal cord slices. (E) Spontaneous EPSC frequency. *P < 0.05, Student’s t-test, n = 5 neurons/group. (F and G) CCI (21 d) increases spontaneous EPSC frequency, which is reversed by the CXCR2 antagonist SB225002 (1 µM). *P < 0.05, Student’s t-test, n = 5 neurons/group.

Article Snippet: The sections were then incubated overnight at 4°C with the following primary antibodies: GFAP antibody (1:1000, mouse; Millipore Bioscience Research Reagents), Cx43 antibody (1:1000, rabbit; Sigma), NeuN antibody (1:1000, mouse; Millipore Bioscience Research Reagents), CXCL1 (1: 200, rabbit; Boster), and CXCR2 antibody (1: 200, rabbit; Boster).

Techniques: Transmission Assay, Western Blot, Control, Injection, Saline, Double Immunostaining, Patch Clamp

Journal: Brain

Article Title: Connexin-43 induces chemokine release from spinal cord astrocytes to maintain late-phase neuropathic pain in mice

doi: 10.1093/brain/awu140

Figure Lengend Snippet: Schematic of working hypothesis for astrocytic Cx43-mediated late-phase neuropathic pain. CCI induces a persistent upregulation of Cx43 in spinal cord astrocytes. Cx43 expression and activity is also upregulated by TNF-α, secreted from microglia. Upregulation of Cx43 hemichannel activities results in CXCL1 release. Astrocytic CXCL1 secretion activates CXCR2 on neurons (central terminals of primary sensory neurons and spinal cord neurons), leading to enhanced excitatory synaptic transmission in nociceptive neurons (e.g. lamina IIo excitatory interneurons) and sustained neuropathic pain in the late-phase. Additionally, CXCL1 can also be secreted from intact or injured primary afferents in the spinal cord especially in the early phase of CCI.

Article Snippet: The sections were then incubated overnight at 4°C with the following primary antibodies: GFAP antibody (1:1000, mouse; Millipore Bioscience Research Reagents), Cx43 antibody (1:1000, rabbit; Sigma), NeuN antibody (1:1000, mouse; Millipore Bioscience Research Reagents), CXCL1 (1: 200, rabbit; Boster), and CXCR2 antibody (1: 200, rabbit; Boster).

Techniques: Expressing, Activity Assay, Transmission Assay

Journal: Redox Report : Communications in Free Radical Research

Article Title: Melittin alleviates sepsis-induced acute kidney injury by promoting GPX4 expression to inhibit ferroptosis

doi: 10.1080/13510002.2023.2290864

Figure Lengend Snippet: Effect of Melittin Attenuates sepsis-AKI. (a) H&E staining of kidney tissues from the Different groups. The red arrows denote the occurrence of tubular dilatation. The blue arrows highlight the swelling of renal tubular epithelial cells. Scale bars: 400 μm. (b) Tubular injury score. (c) Immunohistochemical staining of kidney with primary antibodies against NGAL. The areas marked with red arrows are NGAL-positive regions. Scale bars: 400 μm. (d) Average percentage of positive staining for NGAL per field.(n = 3). (e) Western blotting of NGAL in kidney tissues. (f) Plasma BUN levels. (g) Plasma creatinine levels. * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001; ns, no significant difference between control and LPS group.

Article Snippet: Incubation was performed with the primary antibodies anti-NGAL antibody (PB9609, Boster, China) and anti-4-HNE antibody (AO2685-4, Boster, China).

Techniques: Staining, Immunohistochemical staining, Western Blot, Clinical Proteomics, Control