G-CSF Research Articles

Circulating inflammatory factors and risk causality associated with type 2 diabetic nephropathy: A Mendelian randomization and bioinformatics study.

The main causative factors of diabetic nephropathy (DN), a common complication of diabetes mellitus, are metabolic abnormalities and hemodynamic changes. However, studies have shown that the immune-inflammatory response also plays an important role in DN pathogenesis. Therefore, in this study, we analyzed the causal relationship and immune infiltration between inflammatory factors and DN using Mendelian randomization (MR) and bioinformatics techniques. We analyzed the causal relationship between 91 inflammatory factors and DN using two-sample MR dominated by the results of inverse variance-weighted analysis. Based on the MR analysis, the immune mechanism of inflammatory factors in DN was further explored using immune cell infiltration analysis. MR analysis indicated a positive causal relationship between DN and IL1A, caspase 8 (CASP8), macrophage colony-stimulating factor 1, IL10, STAM-binding protein, and tumor necrosis factor ligand superfamily member 12 (TNFSF12) and a negative causal relationship between DN and cystatin D, fibroblast growth factor 19, neurturin, and TNFSF14. The pathogenic mechanism of CASP8 may involve the recruitment of CD4+ T cells and macrophages for DN infiltration. In this study, we found a causal relationship between DN and IL1A, CASP8, macrophage colony-stimulating factor 1, IL10, STAM-binding protein, TNFSF12, cystatin D, fibroblast growth factor 19, neurturin, and TNFSF14. Bioinformatic immune infiltration analysis further revealed that CASP8 regulates DN by influencing the infiltration of immune cells, such as T cells and macrophages.

Similar Viral and Immune Characteristics of Kaposi Sarcoma in ART-treated People Living With HIV and Older Patients With Classic Kaposi Sarcoma.

Reemergence of human herpesvirus 8 (HHV-8)-induced Kaposi sarcoma (KS) in people living with HIV (PLWH) despite antiretroviral therapy (ART) poses a clinical challenge because they already have favorable CD4 T-cell numbers and undetectable viral loads. We observed that clinical presentation in PLWH on ART resembled classic KS found in older HIV-uninfected patients and hypothesized that immunosenescence may thus play a role in occurrence of KS on ART. We compared viral and immune factors implicated in the development of KS in ART-treated PLWH (HIV KS) and HIV-uninfected classic KS patients (cKS), compared to controls without KS (HIV Control, cControls respectively). Plasma, peripheral blood mononuclear cell, and skin tissues were obtained from 11 HIV KS and 11 cKS patients and 2 groups of age-matched controls. HIV KS participants were younger than cKS (aged 53 vs 75 years). HHV-8 genotypes did not differ between groups. Despite the younger age and a lower CD4/CD8 ratio, activated, exhausted, and senescent T-cell frequencies were similar between HIV KS and cKS. Anti-HHV-8 immunoglobulin G levels were higher and circulating HHV-8 DNA lower in HIV KS compared with cKS. Circulating platelet-derived growth factors AA-BB and granulocyte colony-stimulating factors were higher in HIV KS We observed similar levels of HHV-8 DNA and PD-1 expression in skin lesions from HIV KS and cKS patients. Altogether, early immune senescence could be involved in the development of KS in ART-treated PLWH. Higher anti-HHV-8 immunoglobulin G levels could be linked with lower circulating viral load. Such insights should help developing therapeutical strategies to prevent development and treat KS in PLWH on ART.

Pegylated recombinant human granulocyte colony-stimulating factor for primary prophylaxis of neutropenia in patients with cervical cancer receiving concurrent chemoradiotherapy: a prospective study

BackgroundThis study aimed to investigate the efficacy and safety of pegylated recombinant human granulocyte colony-stimulating factor (PEG-rhG-CSF) for primary prophylaxis of neutropenia in patients with cervical cancer receiving concurrent chemoradiotherapy.MethodsIn this prospective, single-center, single-arm study, we enrolled patients (18–70 years) with 2018 International Federation of Gynecology and Obstetrics (FIGO) stage IIIC1r-IVA and IVB (distant metastasis only with inguinal lymph node metastasis) cervical cancer. Eligible patients should have normal function of the bone marrow (absolute neutrophil count (ANC) ≥ 2.0 × 109/L) and adequate hepatic and renal functions. Key exclusion criteria included: previous chemotherapy and/or radiotherapy; a history of bone marrow dysplasia or other hematopoietic abnormalities. All patients underwent radical radiotherapy (pelvic radiotherapy or extended-field irradiation) plus brachytherapy. The chemotherapy regimen included four cycles of 3-weekly paclitaxel and cisplatin. PEG-rhG-CSF was administered 48–72 h after each treatment cycle. Salvage granulocyte colony-stimulating factor (G-CSF) was only permitted in certain circumstances. The primary endpoint was the incidence of grade 3–4 neutropenia. The secondary endpoints included frequency of febrile neutropenia (FN), chemotherapy completion rate in cycles 2–4, time to complete radiotherapy, and safety.ResultsOverall, 52 patients were enrolled in this study from July 2019 to October 2020. The incidence of grade 3–4 neutropenia was 28.8%, with an average duration of grade 3–4 neutropenia persistence of 3.85 days (1–7 days). The incidence rate of FN was 3.8%. The chemotherapy completion rate was 94.2%, 82.7%, and 75.0% for cycles 2–4, respectively. The incidences of grade 3–4 neutropenia for cycles 1–4 were 9.6% (5/52), 8.2% (4/49), 14.0% (6/43), and 2.6% (1/39), respectively. All patients completed radiotherapy within 8 weeks (median, 48 days; range: 41–56 days), except one patient who withdrew consent and did not receive radiotherapy. Severe non-hematologic toxicity was not observed in any patient.ConclusionPEG-rhG-CSF is an effective and safe prophylactic treatment for neutropenia in patients with cervical cancer undergoing concurrent chemoradiotherapy.Trial registrationChinese Clinical Trial Registry, ChiCTR1900024494. Date of Registration:13/July/2019.

Lenalidomide and pomalidomide modulate hematopoietic cell expansion and differentiation in the presence of MSC

Cytopenia is a well-documented complication in the treatment of hematological malignancies with lenalidomide and pomalidomide. Although prior studies have highlighted direct effects on hematopoietic cells to explain this adverse effect, the involvement of hematopoietic-supportive stroma remains less understood. This study examined the effects of lenalidomide/pomalidomide on the expansion and differentiation of human CD34+ hematopoietic stem/progenitor cells (HSPCs) in vitro, in co-culture with human bone-marrow mesenchymal stromal/stem cells (MSCs). Our findings indicate that lenalidomide/pomalidomide increases the population of immature CD34+CD38− cells while decreasing the number of mature CD34+CD38+ cells, suggesting a mechanism that inhibits early HSPC maturation. This effect persisted across myeloid, megakaryocytic, and erythroid lineages, with MSCs playing a key role in preserving immature progenitors and inhibiting their differentiation. Furthermore, in myeloid differentiation assays augmented by granulocyte-colony stimulating factor, lenalidomide/pomalidomide not only enhanced the presence of CD34+ cells with mature myeloid markers such as CD11b but also reduced the populations lacking CD34 yet positive for these markers, irrespective of MSC presence. Thus, while MSCs support the presence of these immature cell populations, they simultaneously inhibit their maturation. This finding provides novel mechanistic insights into lenalidomide- and pomalidomide-induced cytopenia, and could guide therapeutic strategies for its mitigation.

Sodium-Dependent Glucose Transporter Type 2 Inhibitors as a Breakthrough in Neutropenia and Neutrophil Dysfunction Management in Patients with Glycogen Storage Disease Type Ib

Glycogen storage disease type Ib (GSD Ib) is a rare and extremely severe disease included in the group of hereditary carbohydrate metabolism disorders. The condition is caused by pathogenic variants in the SLC37A4 gene leading to glucose metabolic disorder in the liver and kidneys, and as a result to severe organomegaly, hypoglycemia, and metabolic decompensation. Moreover, neutropenia and neutrophil dysfunction are noted in patients with GSD Ib. The use of granulocyte colony stimulating factor only increases the number of dysfunctional neutrophils without affecting their functional activity, what determines the inefficacy of such treatment. In recent years, the mechanism of neutropenia in GSD Ib has been clarified, so new therapeutic agents for its relief have been created. This article presents the overview of data on the successful results of renal sodium-glucose cotransporter type 2 inhibitors (gliflozins) usage in patients with GSD Ib.

Serum GM-CSF level is a predictor of treatment response to tocilizumab in rheumatoid arthritis patients: a prospective observational cohort study

BackgroundThe aim of this prospective observational cohort study was to unveil the predictors of treatment response to tocilizumab (TCZ) therapy in rheumatoid arthritis (RA) patients, in terms of clinical characteristics and serum proinflammatory cytokines, especially to explore the predictive value of granulocyte macrophage-colony stimulating factor (GM-CSF).MethodsActive adult RA patients with inadequate response to MTX intending to receive TCZ therapy were recruited prospectively in the study. A total of 174 severe RA patients were included for the identification of the associations between treatment response and the following characteristic features: demographics, medications, disease activity, serum proinflammatory cytokines and so on.ResultsDisease duration (OR = 0.996), tender joint count (TJC)/68 (OR = 0.943), neutrophil ratio (W4/baseline) (OR = 0.224), the high level of GM-CSF > 5 ng/ml (OR = 0.414) at baseline were the independent adverse predictors of good response assessed by clinical disease activity index (CDAI) at week 24 (W24) for TCZ therapy in RA patients. Moreover, DAS28-ESR (OR = 2.951, P = 0.002) and the high level of GM-CSF > 10 ng/ml at baseline (OR = 5.419, P = 0.002) were independent predictors of poor response, but not the high level of GM-CSF > 5 ng/ml (OR = 2.713, P = 0.054). The patients in the high GM-CSF group had significantly higher DAS28-ESR and serum levels of cytokines (IL-17A, IL-1β, IL-6, TNF-α) at baseline, as well as significantly higher rate of non-good response (62.8% vs. 39.4%, P = 0.010) and poor response (27.9% vs. 9.1%, P = 0.004) than the low GM-CSF group at W24. In addition, poor responders had significantly higher levels of GM-CSF with concomitant increase in the serum levels of IL-17A and IL-1β at baseline than those in moderate and good response groups, while serum levels of IL-6 and TNF-α at baseline were not significantly different in three response groups.ConclusionThe high levels of GM-CSF (> 5 ng/ml and > 10 ng/ml) at baseline were the independent predictors of non-good response and poor response to TCZ at W24 respectively. The high level of GM-CSF at baseline is a marker of high disease activity and a predictor of poor response to TCZ in severe RA patients, which may facilitate the development of individualized treatment strategies for refractory RA.

Focal Adhesion Kinase and Colony Stimulating Factors: Intestinal Homeostasis and Innate Immunity Crosstalk.

Thousands struggle with acute and chronic intestinal injury due to various causes. Epithelial intestinal healing is dependent on phenotypic transitions to a mobile phenotype. Focal adhesion kinase (FAK) is a ubiquitous protein that is essential for cell mobility. This phenotype change is mediated by FAK activation and proves to be a promising target for pharmaceutical intervention. While FAK is crucial for intestinal healing, new evidence connects FAK with innate immunity and the importance it plays in macrophage/monocyte chemotaxis, as well as other intracellular signaling cascades. These cascades play a part in macrophage/monocyte polarization, maturation, and inflammation that is associated with intestinal injury. Colony stimulating factors (CSFs) such as macrophage colony stimulating factor (M-CSF/CSF-1) and granulocyte macrophage colony stimulating factor (GM-CSF/CSF-2) play a critical role in maintaining homeostasis within intestinal mucosa by crosstalk capabilities between macrophages and epithelial cells. The communication between these cells is imperative in orchestrating healing upon injury. Diving deeper into these connections may allow us a greater insight into the role that our immune system plays in healing, as well as a better comprehension of inflammatory diseases of the gut.

Combinatorial macrophage induced innate immunotherapy against Ewing sarcoma: Turning “Two Keys” simultaneously

BackgroundMacrophages play important roles in phagocytosing tumor cells. However, tumors escape macrophage phagocytosis in part through the expression of anti-phagocytic signals, most commonly CD47. In Ewing sarcoma (ES), we found that tumor cells utilize dual mechanisms to evade macrophage clearance by simultaneously over-expressing CD47 and down-regulating cell surface calreticulin (csCRT), the pro-phagocytic signal. Here, we investigate the combination of a CD47 blockade (magrolimab, MAG) to inhibit the anti-phagocytic signal and a chemotherapy regimen (doxorubicin, DOX) to enhance the pro-phagocytic signal to induce macrophage phagocytosis of ES cells in vitro and inhibit tumor growth and metastasis in vivo.MethodsMacrophages were derived from human peripheral blood monocytes by granulocyte–macrophage colony-stimulating factor (GM-CSF) and macrophage colony-stimulating factor (M-CSF). Flow cytometry- and microscopy-based in-vitro phagocytosis assays were performed to evaluate macrophage phagocytosis of ES cells. Annexin-V assay was performed to evaluate apoptosis. CD47 was knocked out by CRISPR/Cas9 approach. ES cell-based and patient-derived-xenograft (PDX)-based mouse models were utilized to assess the effects of MAG and/or DOX on ES tumor development and animal survival. RNA-Seq combined with CIBERSORTx analysis was utilized to identify changes in tumor cell transcriptome and tumor infiltrating immune cell profiling in MAG and/or DOX treated xenograft tumors.ResultsWe found that MAG significantly increased macrophage phagocytosis of ES cells in vitro (p < 0.01) and had significant effect on reducing tumor burden (p < 0.01) and increasing survival in NSG mouse model (p < 0.001). The csCRT level on ES cells was significantly enhanced by DOX in a dose- and time-dependent manner (p < 0.01). Importantly, DOX combined with MAG significantly enhanced macrophage phagocytosis of ES cells in vitro (p < 0.01) and significantly decreased tumor burden (p < 0.01) and lung metastasis (p < 0.0001) and extended animal survival in vivo in two different mouse models of ES (p < 0.0001). Furthermore, we identified CD38, CD209, CD163 and CD206 as potential markers for ES-phagocytic macrophages. Moreover, we found increased M2 macrophage infiltration and decreased expression of Cd209 in the tumor microenvironment of MAG and DOX combinatorial therapy treated tumors.ConclusionsBy turning “two keys” simultaneously to reactivate macrophage phagocytic activity, our data demonstrated an effective and highly translatable alternative therapeutic approach utilizing innate (tumor associated macrophages) immunotherapy against high-risk metastatic ES.

Association between immune cells, inflammatory cytokines, and sarcopenia: Insights from a Mendelian randomization analysis

BackgroundRecent studies have suggested a possible link between sarcopenia, immune dysregulation, and chronic inflammation, although the specific immune components implicated remain unclear. This investigation employs Mendelian Randomization (MR) to explore the reciprocal relationship between immune cells, inflammatory markers, and sarcopenia. MethodWe performed two-sample and multivariate MR analyses using publicly accessible genome-wide association studies (GWAS) summary statistics. Our analyses included 731 immune cells, 41 inflammatory cytokines, and sarcopenia related traits (appendicular lean mass [ALM], low hand-grip strength [LHS], and walking pace [WP]), with additional sensitivity analyses conducted to confirm the findings. ResultsAfter false discovery rate (FDR) correction, significant associations were found between ten immune traits and ALM, with the CD127 marker in the Treg panel showing consistent positive correlation across four sites. In contrast, NKT%lymphocyte negatively correlated with WP (OR = 0.99, P = 0.023). In terms of inflammatory cytokines, macrophage colony-stimulating factor (MCSF) (OR = 1.03, P = 0.024) and hepatocyte growth factor (HGF) (OR = 1.03, P = 0.002) demonstrated positive associations with ALM, while interleukin-16 (IL-16) (OR = 0.99, P = 0.006) was inversely related. The reverse Mendelian randomization analysis found no direct causal links between sarcopenia traits and immune or inflammatory markers. Sensitivity analyses underscored the findings' resilience to pleiotropy, and adjusting for inter-trait dynamics weakened these relationships in the multivariable MR analysis. ConclusionOur study reveals causal associations between specific immune phenotypes, inflammatory cytokines, and sarcopenia, providing insight into the development of sarcopenia and potential treatment strategies.

Single-Cell RNA Sequencing Reveals Immunomodulatory Effects of Stem Cell Factor and Granulocyte Colony-Stimulating Factor Treatment in the Brains of Aged APP/PS1 Mice.

Alzheimer's disease (AD) leads to progressive neurodegeneration and dementia. AD primarily affects older adults with neuropathological changes including amyloid-beta (Aβ) deposition, neuroinflammation, and neurodegeneration. We have previously demonstrated that systemic treatment with combined stem cell factor (SCF) and granulocyte colony-stimulating factor (G-CSF) (SCF+G-CSF) reduces the Aβ load, increases Aβ uptake by activated microglia and macrophages, reduces neuroinflammation, and restores dendrites and synapses in the brains of aged APPswe/PS1dE9 (APP/PS1) mice. However, the mechanisms underlying SCF+G-CSF-enhanced brain repair in aged APP/PS1 mice remain unclear. This study used a transcriptomic approach to identify the potential mechanisms by which SCF+G-CSF treatment modulates microglia and peripheral myeloid cells to mitigate AD pathology in the aged brain. After injections of SCF+G-CSF for 5 consecutive days, single-cell RNA sequencing was performed on CD11b+ cells isolated from the brains of 28-month-old APP/PS1 mice. The vast majority of cell clusters aligned with transcriptional profiles of microglia in various activation states. However, SCF+G-CSF treatment dramatically increased a cell population showing upregulation of marker genes related to peripheral myeloid cells. Flow cytometry data also revealed an SCF+G-CSF-induced increase of cerebral CD45high/CD11b+ active phagocytes. SCF+G-CSF treatment robustly increased the transcription of genes implicated in immune cell activation, including gene sets that regulate inflammatory processes and cell migration. The expression of S100a8 and S100a9 was robustly enhanced following SCF+G-CSF treatment in all CD11b+ cell clusters. Moreover, the topmost genes differentially expressed with SCF+G-CSF treatment were largely upregulated in S100a8/9-positive cells, suggesting a well-conserved transcriptional profile related to SCF+G-CSF treatment in resident and peripherally derived CD11b+ immune cells. This S100a8/9-associated transcriptional profile contained notable genes related to pro-inflammatory and anti-inflammatory responses, neuroprotection, and Aβ plaque inhibition or clearance. Altogether, this study reveals the immunomodulatory effects of SCF+G-CSF treatment in the aged brain with AD pathology, which will guide future studies to further uncover the therapeutic mechanisms.

Rhizoma Zedoariae Inhibits G-CSF-Induced MDSCs-like Differentiation of THP-1 Cells

Objective: Rhizoma Zedoariae is widely used in the treatment of solid tumors in China, but there has been no deep research on its role in the formation of timmunosuppressive environments mediated by myeloid-derived suppressor cells (MDSCs) in breast cancer. This research aims to investigate the effect of Rhizoma Zedoariae extract and its active compounds curcumin and germacrone on MDSCs-like differentiation by a novel Granulocyte colony-stimulating factor (G-CSF) induced THP-1 cell model. Methods: THP-1 cells were treated with Rhizoma Zedoariae extract and its active compounds (curcumin, germacrone, curcumol, curcumenol, and curdione) at different concentrations (0.01, 0.1, 0.3, 1, 10, 30, 100, and 300 μg/mL), followed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay to detect the changes of cell viability in THP-1. Quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR) was performed to quantify the mRNA expression levels of transforming growth factor-β (TGF-β), Interleukin-10 (IL-10), Interleukin-6 (IL-6), Janus kinase 1 (JAK1), nuclear factor kappa-B (NF-κB), signal transducer and activator of transcription 3 (STAT3) and arginase-1 (ARG-1), to detected the differentiation of THP-1 into MDSCs-like cells. Results: G-CSF can induce the cell viability and MDSCs-like differentiation of THP-1 cells. Rhizoma Zedoariae extract (0.1, 1, 10, 30, 100, 300 μg/mL), germacrone (0.1, 1, 10, 30, 100, 300 μg/mL) and curcumin (0.01, 0.1, 1, 10, 30, 100 μg/mL) exerted obvious inhibitory effects (&gt;70%) on THP-1 cell viability. G-CSF increased the ARG-1 and IL-10 mRNA levels. Rhizoma Zedoariae extract in 100 μg/mL decreased the ARG-1 and IL-10 mRNA levels. Furthermore, Rhizoma Zedoariae extract in 300 μg/mL also significantly decreased the mRNA expressions of JAK1 and STAT3 (p &lt; 0.001). Conclusion: Rhizoma Zedoariae extract maybe inhibit G-CSF-induced MDSCs-like differentiation of THP-1 through the STAT3 pathway. The active compound that exerts this effect may be germacrone and curcumin.

Use of letermovir for cytomegalovirus primary prophylaxis in lung transplant recipients.

Cytomegalovirus (CMV) is a driver of negative outcomes after lung transplant (LTX) and primary prophylaxis (PPX) with valganciclovir (VGC) is standard-of-care. VGC is associated with myelosuppression, prompting interest in letermovir (LTV). Adults receiving LTX between April 1, 2015, and July 30, 2022, at our institution were evaluated. Patients were excluded if low CMV risk (D-/R-), survived <90 days post-LTX, or transferred care before PPX withdrawal. Primary outcomes were leukopenia (white blood cell count [WBC] ≤ 3.0×109/L), severe leukopenia (WBC ≤ 2.0×109/L), and neutropenia (absolute neutrophil count ≤ 1500 cells/µL) requiring granulocyte-colony stimulating factor (GCSF) on PPX. Secondary outcomes included breakthrough CMV infection and post-PPX CMV infection. 204 patients met inclusion criteria: 175 patients on VGC and 29 patients on LTV (after VGC conversion). Most patients received bilateral LTX (62.7%) with non-lymphocyte-depleting induction (96.6%) and moderate-risk serostatus (D+/R+, 48.5%). Patients transitioned from VGC to LTV after a mean of 178 days (SD 80.8 days) post-transplant. Patients on VGC experienced significantly more leukopenia (82.3%vs. 58.6%, p = 0.008), severe leukopenia (57.1%vs. 31.0%, p = 0.016), and neutropenia requiring GCSF (70.9%vs. 51.7%, p = 0.048). Breakthrough (5.7%vs. 3.4%, p = 0.955) and post-PPX (24.6%vs. 37.9%, p = 0.199) infections were similar. A subgroup analysis of patients with high-risk serostatus showed similar trends, though did not reach statistical significance. In this single-center study, the incidence of leukopenia and neutropenia requiring GCSF were reduced with LTV compared to VGC. Breakthrough and post-PPX infections were not significantly different. This evidence suggests that LTV has comparable efficacy with reduced myelosuppression compared to VGC in LTX recipients, and may be an appropriate alternative for PPX.

New mechanistic understanding of osteoclast differentiation and bone resorption mediated by P2X7 receptors and PI3K-Akt-GSK3β signaling

ObjectiveOsteoporosis is a global health issue characterized by decreased bone mass and microstructural degradation, leading to an increased risk of fractures. This study aims to explore the molecular mechanism by which P2X7 receptors influence osteoclast formation and bone resorption through the PI3K-Akt-GSK3β signaling pathway.MethodsAn osteoporosis mouse model was generated through ovariectomy (OVX) in normal C57BL/6 and P2X7f/f; LysM-cre mice. Osteoclasts were isolated for transcriptomic analysis, and differentially expressed genes were selected for functional enrichment analysis. Metabolite analysis was performed using liquid chromatography-tandem mass spectrometry (LC–MS/MS), and multivariate statistical analysis and pattern recognition were used to identify differential lipid metabolism markers and their distribution. Bioinformatics analyses were conducted using the Encyclopedia of Genes and Genomes database and the MetaboAnalyst database to assess potential biomarkers and create a metabolic pathway map. Osteoclast precursor cells were used for in vitro cell experiments, evaluating cell viability and proliferation using the Cell Counting Kit 8 (CCK-8) assay. Osteoclast precursor cells were induced to differentiate into osteoclasts using macrophage colony-stimulating factor (M-CSF) and receptor activator of nuclear factor kappa-beta ligand (RANKL), and tartrate-resistant acid phosphatase (TRAP) staining was performed to compare differentiation morphology, size, and quantity between different groups. Western blot analysis was used to assess the expression of differentiation markers, fusion gene markers, and bone resorption ability markers in osteoclasts. Immunofluorescence staining was employed to examine the spatial distribution and quantity of osteoclast cell skeletons, P2X7 protein, and cell nuclei, while pit assay was used to evaluate osteoclast bone resorption ability. Finally, in vivo animal experiments, including micro computed tomography (micro-CT), hematoxylin and eosin (HE) staining, TRAP staining, and immunohistochemistry, were conducted to observe bone tissue morphology, osteoclast differentiation, and the phosphorylation level of the PI3K-Akt-GSK3β signaling pathway.ResultsTranscriptomic and metabolomic data collectively reveal that the P2X7 receptor can impact the pathogenesis of osteoporosis through the PI3K-Akt-GSK3β signaling pathway. Subsequent in vitro experiments showed that cells in the Sh-P2X7 + Recilisib group exhibited increased proliferative activity (1.15 versus 0.59), higher absorbance levels (0.68 versus 0.34), and a significant increase in resorption pit area (13.94 versus 3.50). Expression levels of osteoclast differentiation-related proteins MMP-9, CK, and NFATc1 were markedly elevated (MMP-9: 1.72 versus 0.96; CK: 2.54 versus 0.95; NFATc1: 3.05 versus 0.95), along with increased fluorescent intensity of F-actin rings. In contrast, the OE-P2X7 + LY294002 group showed decreased proliferative activity (0.64 versus 1.29), reduced absorbance (0.34 versus 0.82), and a significant decrease in resorption pit area (5.01 versus 14.96), accompanied by weakened expression of MMP-9, CK, and NFATc1 (MMP-9: 1.14 versus 1.79; CK: 1.26 versus 2.75; NFATc1: 1.17 versus 2.90) and decreased F-actin fluorescent intensity. Furthermore, in vivo animal experiments demonstrated that compared with the wild type (WT) + Sham group, mice in the WT + OVX group exhibited significantly increased levels of CTX and NTX in serum (CTX: 587.17 versus 129.33; NTX: 386.00 versus 98.83), a notable decrease in calcium deposition (19.67 versus 53.83), significant reduction in bone density, increased trabecular separation, and lowered bone mineral density (BMD). When compared with the KO + OVX group, mice in the KO + OVX + recilisib group showed a substantial increase in CTX and NTX levels in serum (CTX: 503.50 versus 209.83; NTX: 339.83 versus 127.00), further reduction in calcium deposition (29.67 versus 45.33), as well as decreased bone density, increased trabecular separation, and reduced BMD.ConclusionP2X7 receptors positively regulate osteoclast formation and bone resorption by activating the PI3K-Akt-GSK3β signaling pathway.

Multiple small-dose infusions of G-CSF-mobilized haploidentical lymphocytes after autologous haematopoietic stem cell transplantation for acute myeloid leukaemia.

This retrospective study analysed 106 acute myeloid leukaemia (AML) patients undergoing autologous haematopoietic stem cell transplantation (ASCT) to assess the impact of multiple small-dose infusions of granulocyte-colony-stimulating factor (G-CSF)-mobilized haploidentical lymphocytes as post-ASCT maintenance therapy. Among them, 50 patients received lymphocyte maintenance therapy, 21 received alternative maintenance therapy, and 35 received no maintenance therapy. Patients receiving lymphocyte maintenance therapy demonstrated significantly higher overall survival (OS) and disease-free survival (DFS) compared to those without maintenance therapy, with 4-year OS and DFS rates notably elevated. While there were no significant differences in recurrence rates among the three groups, lymphocyte maintenance therapy showcased particular benefits for intermediate-risk AML patients, yielding significantly higher OS and DFS rates and lower relapse rates compared to alternative maintenance therapy and no maintenance therapy. The study suggests that multiple small-dose infusions of G-CSF-mobilized haploidentical lymphocytes may offer promising outcomes for AML patients after ASCT, particularly for those classified as intermediate-risk. These findings underscore the potential efficacy of lymphocyte maintenance therapy in reducing disease relapse and improving long-term prognosis in this patient population.

Moxibustion combined with guasha therapy for recurrent neutropenia following multiple cycles of chemotherapy of ovarian cancer: A case report

Neutropenia, a common side effect of chemotherapy for ovarian cancer, was observed in a 47-year-old female patient undergoing a six-cycle chemotherapy regimen. She experienced recurrent neutropenia and leukopenia but refused granulocyte colony-stimulating factor (G-CSF) due to severe bone pain and high costs. Moxibustion combined with guasha therapy (MGT) was administered each time neutropenia occurred. The treatment involved guasha therapy on the bladder meridian (BL) and the governor vessel (GV), followed by moxibustion at Zhongwan (CV 12), Guanyuan (CV 4), and Shenzhu (GV 12) points over 2–3 days. This approach led to the recovery of neutrophil and leukocyte counts, enabling the patient to complete six chemotherapy cycles without G-CSF. These findings suggest that MGT may enhance neutrophil and leukocyte counts in patients with chemotherapy-induced myelosuppression, presenting a potential alternative for those intolerant to G-CSF. However, further high-quality research is needed to confirm its efficacy.

Addition of GM-CSF during in vitro oocyte maturation improves embryo development and implantation and birth rate in mice.

The present study determined whether adding granulocyte macrophage colony stimulating factor (GM-CSF) during in vitro oocyte maturation (IVM) could improve oocyte developmental competence by examining embryo development and implantation and birth rates following embryo transfer in mice. In an initial dose response experiment, we demonstrated that the addition of 2 and 10 ng/mL of GM-CSF during IVM increased cumulus expansion (P<0.05) but did not affect fertilisation rate compared with the control group. The addition of 10 ng/mL increased blastocyst rate (17.0%; P<0.05) and tended to increase the number of good quality blastocysts present at 96 h of culture (+19.4%; P=0.06) and increased blastocyst inner cell mass (+25.2%; P<0.001), trophectoderm (+29.9%; P<0.01), and total cell numbers (+28.6%; P<0.05). GM-CSF also reduced the incidence of DNA damage in blastocysts in the 10 ng/mL group (-16.2%) compared with the control group. These improvements translated into increases in implantation rate (+21.0%; P<0.05) and birth rate (+17.0%; P<0.05) following the transfer of vitrified blastocysts.GM-CSF treatment did not alter any fetal and placental parameters. Together these results suggest that the addition of GM-CSF during IVM may improve livestock in vitro embryo production and human IVM.

Optimized high-dose granulocyte transfusions for the treatment of infections in neutropenic patients: A single-center retrospective analysis.

Granulocyte transfusions for patients with prolonged neutropenia and severe infections has been a controversial practice. Previous studies suggest a benefit of high-dose granulocyte transfusions (≥0.6 × 109/kg), although, until recently, the consistent production of high-dose units has been challenging. Here, we present our experience and results utilizing high-dose granulocyte transfusions at a large, tertiary academic medical center for the treatment of infections in adult, neutropenic patients. A retrospective chart review (2018-2021) was conducted for all patients who received high-dose granulocyte transfusions from donors stimulated with granulocyte colony-stimulating factor (G-CSF) and dexamethasone. Gathered parameters included patient demographics, clinical history, infection status, dose, clinical outcomes, pre- and post-absolute neutrophil count (ANC), and transfusion times including time between granulocyte collection, administration, and posttransfusion ANC count. Gathered parameters were summarized using descriptive statistics, outcomes were assessed utilizing Kaplan-Meier curves/log-rank/regression testing. Totally 28 adult, neutropenic patients refractory to antimicrobial agents and/or G-CSF received a total of 173 granulocyte concentrates. Median ANC increased from 0.7 × 109/L pre-transfusion to 1.6 × 109/L posttransfusion. The mean granulocyte yield was 77.4 × 109 resulting in an average dose per kilogram of 0.90 × 109 ± 0.30 × 109 granulocytes. Composite day 42 survival and microbial response was 42.9% (n = 12/28) without significant adverse reactions. Here, we demonstrate the successful and safe implementation of high-dose granulocyte transfusions for neutropenic patients. Given the rapid and consistent production, distribution, and improved granulocyte quality, further investigations to determine the clinical efficacy of G-CSF primed granulocyte transfusions is now possible.

O-076 Novel method to enhance preantral follicle growth by granulocyte colony-stimulating factor administration improves embryos and pregnancy rate in poor ovarian reserve: a randomized controlled trial

Abstract Study question Whether priming with granulocyte colony-stimulating factor (G-CSF) before ART improves embryo and pregnancy rate while increasing anti-Müllerian hormone (AMH) in patients with poor ovarian reserve. Summary answer G-CSF priming improved embryonic development and pregnancy rate in ART with poor ovarian reserve, simultaneously increasing serum AMH. Enhanced preantral follicle growth likely was responsible. What is known already Eleven years ago, we treated repeated implantation failure in 10 women with diminished ovarian reserve by administering G-CSF with embryo transfers. No implantation succeeded directly, but serendipitously 3 of the 10 spontaneously became pregnant 2 months later. One of these pregnancies involved twins in a 45-year-old woman without ovulation induction. We hypothesized that G-CSF administration might have stimulated preantral follicle growth, with improved ovulation after 2 months. In animal models with diminished ovarian reserve, G-CSF administration consistently increased ovarian preantral follicles and AMH. Therefore, we designed this clinical trial. Study design, size, duration Patients were prospectively randomized to priming or control groups. All patients initially underwent conventional infertility treatment for 2 consecutive cycles in which the priming group but not controls received a subcutaneous G-CSF priming injection during the early luteal phase. Each group then underwent 1 cycle of IVF/ICSI and fresh embryo transfer (IVF/ICSI-fresh ET), followed by cryopreserved ET if needed until live birth or embryo depletion. AMH was measured before and after priming. Participants/materials, setting, methods Hundred ART patients 20 to 42 years old with AMH below 2 ng/mL were randomized to priming or control groups (50 patients each). None had over 1 ART failure, day-3 follicle-stimulating hormone (FSH) above 30 IU/L, uterine anomalies, or a partner with azoospermia. Main results and the role of chance Fertilization rate, embryonic development, and implantation rate by fresh ET were significantly improved by G-CSF priming. Clinical and ongoing pregnancy rates by IVF/ICSI-fresh ET were significantly higher with priming (30% and 26% in 47 ART patients; 3 delivered with conventional treatment) than in controls (12% and 10% in 49 ART patients; 1 dropped out). With priming, significantly more patients achieved cryopreservation of redundant blastocysts (53% with priming vs. 24% in controls). The cumulative live birth rate was 32% in 50 patients with priming, significantly higher than 14% in 49 controls (relative risk, 2.8; 95% confidence interval, 1.04-7.7). Infants derived from priming had no congenital anomalies, while infant weights, birth weeks, and Apgar scores were similar between groups. Among 4 variables (age, day-3 FSH, AMH, and priming), logistic regression significantly associated age and priming with cumulative live birth. Priming significantly increased serum AMH. The efficacy of G-CSF priming was not associated with genotypes for killer-cell immunoglobulin-like receptors. No adverse effects of priming were observed. Limitations, reasons for caution Enhancement of preantral follicle growth by G-CSF administration needs to be confirmed histologically as this clinical trial could not allow ovarian biopsies for its purpose of infertility treatments. Wider implications of the findings This study proposes a novel, simple, and safe treatment for poor ovarian reserve. In addition, enhancement of preantral follicle growth by G-CSF priming may also improve ovulatory dysfunctions in polycystic ovary syndrome with elevated AMH, reflecting pooling of retarded/arrested growing preantral follicles. Trial registration number UMIN000013956

Demonstration of Physicochemical and Functional Similarity of Biosimilar Pegfilgrastim-cbqv to Pegfilgrastim.

Pegfilgrastim-cbqv/CHS-1701 (UDENYCA®) (hereafter referred to as pegfilgrastim-cbqv) was approved in 2018 by the US Food and Drug Administration as a biosimilar for pegfilgrastim (Neulasta®) (hereafter referred to as pegfilgrastim). Both pegfilgrastim-cbqv and pegfilgrastim are conjugates of recombinant human granulocyte colony stimulating factor (r-metHuG-CSF) with a 20kDa polyethylene glycol (PEG) indicated to decrease the incidence of infection, as manifested by febrile neutropenia, in patients receiving myelosuppressive anticancer drugs. The demonstration of analytical similarity for PEG-protein conjugates presents unique challenges since both the protein and PEG attributes must be characterized. The current study demonstrates the analytical similarity of pegfilgrastim-cbqv and the reference product, pegfilgrastim. In addition to the physicochemical and functional characterization of the protein, the study assessed attributes specific to PEGylation including PEG size and polydispersity, site of attachment, linker composition, and PEGylation process-related variants. The structural, functional, and stability attributes of pegfilgrastim-cbqv and pegfilgrastim were compared using state-of-the-art analytical methods. For the protein, the primary structure, disulfide structure, and secondary and tertiary structures were assessed using traditional protein characterization techniques such as mass spectrometry (MS), circular dichroism (CD), intrinsic fluorescence, and differential scanning calorimetry (DSC), as well as more advanced techniques such as two-dimensional (2D) nuclear magnetic resonance (NMR) and hydrogen deuterium exchange (HDX). For the PEG moiety, the site of attachment, occupancy, linker composition, size and polydispersity were compared using mass spectrometry (both intact and after endoprotease digestion), multiangle light scattering detection (MALS), and Edman degradation. Purity assessments included the assessment of both protein variants and PEGylation variants using chromatographic and electrophoretic analytical separation techniques. The functional similarity between pegfilgrastim-cbqv and pegfilgrastim was compared using both a cell-based bioassay and surface plasmon resonance (SPR). The degradation rates and stability profiles were compared under accelerated and stressed conditions. Biosimilarity was demonstrated by a thorough assessment of physiochemical and functional attributes, as well as comparative stability, of pegfilgrastim-cbqv relative to pegfilgrastim. These studies demonstrated identical primary structure and disulfide structure, highly similar secondary and tertiary structure, as well as functional similarity. The impurity profile of pegfilgrastim-cbqv was comparable to that of pegfilgrastim with only minor differences in PEGylation variants and a slight offset in the PEG molar mass. These differences were not clinically relevant. The degradation profiles were qualitatively and quantitatively similar under accelerated and stress conditions. The structural, functional, and stability data demonstrate that pegfilgrastim-cbqv is highly similar to the reference product, pegfilgrastim.

P-803 Intra-ovarian injection of plasma rich in stem cell-secreted and platelet-enclosed factors to promote follicle rescue and reproductive outcomes in women with very low ovarian reserve

Abstract Study question Could intraovarian injection of plasma rich in stem-cell and platelet growth factors optimize ovarian reserve and IVF outcomes in women with very low ovarian reserve? Summary answer Intraovarian injection of G-CSF mobilized plasma rich in grown factors improved follicle count since first follow-up and led to better fecundation, implantation and pregnancy’ rates. What is known already Autologous stem cell ovarian transplantation (ASCOT) of bone marrow derived stem cells optimized ovarian reserve and reproductive outcomes in poor responders (POR) and this effect seems to be due to the growth factors secreted by the stem cells, acting through paracrine pathways. Similarly, platelet-rich Plasma (PRP) has been also offered to promote ovarian reactivation in patients with impaired ovarian reserve by providing positive growth factors contained in platelets. Indeed, a technique that combines both the non-cellular components of ASCOT and PRP (ASCOT-1) has been proposed to optimize follicle rescue and IVF outcomes in women with very low reproductive prognosis. Study design, size, duration Retrospective observational study with 159 women (≤ 45 years) diagnosed with POR and primary ovarian insufficiency (POI) according to ESHRE criteria, who underwent the ASCOT-1 technique after refusing oocyte donation, between June 2021-October 2023 at IVIRMA Alicante, Spain (IRB approval 2310-FIVI-188-SH). The ASCOT-1 consisted in the intraovarian injection of plasma rich in both stem cell-secreted and platelet-enclosed factors obtained after granulocyte colony stimulating factor (G-CSF) mobilization and PRP obtention (Endoret Kit BTI, Biotechnology institute). Participants/materials, setting, methods Medical records of patients who underwent the ASCOT-1 as an ovarian reactivation treatment were examined. Follow up of AFC and AMH was carried out for a maximum of 4 menstrual cycles to evaluate follicular activation. Moreover, when follicle growth was detected, patients underwent a controlled ovarian stimulation (COS). IVF and reproductive outcomes were compared with previous COS cycles started in our clinics, when available. T-test was used where appropriate. Main results and the role of chance Overall 159 POR/POI women (age 38.7±4.2years, AMH0 0.22±0.41ng/ml; AFC0 1.4±1.9) underwent the ASCOT-1 technique. Mean volume of plasma injected was 6.8±0.8ml, distributed equally in both ovaries. First follow-up was done in 131 women showing statistically significant improvements in the AFC during the entire follow-up compared to the basal levels measured before the treatment (AFC1 3.3±3.4, p &amp;lt; 0.001; AFC2 3.3±3.6, p &amp;lt; 0.001; AFC3 2.9±2.6, p &amp;lt; 0.001; AFC4 2.4±2.6, p = 0.07). After ASCOT-1, 98 COS were initiated in 48 women in our clinics, which were compared to their previous 66 cycles. Although age was significantly higher in their posterior cycles (38.0±2.9 vs40.0±3.6; p &amp;lt; 0.001), after ASCOT-1 oocyte retrieval was reached in a greater amount of cases (75.5% vs87.5%, p=NS). ASCOT-1 showed a positive, but non statistically significant effect in the number of MII-oocytes (2.8±4.1 vs3.2±3.9, p=NS) and blastocysts obtained (1.8±2.8 vs2.4±3.1, p=NS), but relevant improvements were observed in quality related parameters such as oocyte maturation (61.5%, 95%IC 45.8-77.3, vs86.1%, 95I%IC 69.9-85.5; p 0.03), fertilization (62.6%, 95%CI 53.0-71.3, vs76.5%, 95%CI 70.0-82.0; p &amp;lt; 0.001), implantation (25.0%, 95%CI 8.3-52.5 vs41.1%, 95%CI 19.4-66.5; p &amp;lt; 0.001) and clinical pregnancy rate (25.0%, 95%CI 8.3-52.5 vs46.6%, 95%CI 22.2-72.5; p &amp;lt; 0.001). Ultimately, 14 clinical pregnancies (7 after ET and 7 by natural conception) were obtained after ASCOT-1. Limitations, reasons for caution This retrospective study involves short periods of follow up and limited amount of IVF attempts. Thus, further prospective studies with increased sample size are required to validate our results, to establish the duration of the effects, and to identify which subgroups of patients could benefit the most from this treatment. Wider implications of the findings Growth factors contained in the G-CSF mobilized PRP could play a role in follicular development, providing a new tool to increase the reproductive potential of women with very low ovarian reserve where oocyte donation is the only practical option, allowing them to access to homologous IVF treatment. Trial registration number 2310-FIVI-188-SH