[Three-dimensional finite element analysis of lumbar disc herniation under different body positions].

OBJECTIVE: To campare biomechanical effects of different postural compression techniques on three-dimensional model of lumbar disc herniation (LDH) by finite element analysis. METHODS: Lumbar CT image of a 48-year-old female patient with LDH (heighted 163 cm, weighted 53 kg) was collected. Mimics 20.0, Geomagic Studio, Solidwords and other software were used to establish three-dimensional finite element model of LDH on L4,5 segments. Compression techniques under horizontal position, 30° forward bending and 10° backward extension were simulated respectively. After applying the pressure, the effects of compression techniques under different positions on stress, strain and displacement of various tissues of intervertebral disc and nerve root were observed. RESULTS: L4, 5 segment finite element model was successfully established, and the model was validated. When compression manipulation was performed on the horizontal position, 30° flexion and 10° extension, the annular stress were 0.732, 5.929, 1.286 MPa, the nucleus pulposus stress were 0.190, 1.527, 0.295 MPa, and the annular strain were 0.097, 0.922 and 0.424, the strain sizes of nucleus pulposus were 0.153, 1.222 and 0.282, respectively. The overall displacement distance of intervertebral disc on Y direction were -3.707, -18.990, -4.171 mm, and displacement distance of nerve root on Y direction were +7.836, +5.341, +3.859 mm, respectively. The relative displacement distances of nerve root and intervertebral disc on Y direction were 11.543, 24.331 and 8.030 mm, respectively. CONCLUSION: Compression manipulation could make herniated intervertebral disc produce contraction and retraction trend, by increasing the distance between herniated intervertebral disc and nerve root, to reduce symptoms of nerve compression, to achieve purpose of treatment for patients with LDH, in which the compression manipulation is more effective when the forward flexion is 30°.

High and selective cytotoxicity of ex vivo expanded allogeneic human natural killer cells from peripheral blood against bladder cancer: implications for natural killer cell instillation after transurethral resection of bladder tumor.

BACKGROUND: Non-muscle-invasive bladder cancer (NMIBC) is treated with transurethral resection of bladder tumor (TURBT) followed by intravesical instillation of chemotherapy or Bacillus Calmette-Guérin therapy. However, these treatments have a high recurrence rate and side effects, emphasizing the need for alternative instillations. Previously, we revealed that expanded allogeneic human natural killer (NK) cells from peripheral blood are a promising cellular therapy for prostate cancer. However, whether NK cells exhibit a similar killing effect in bladder cancer (BCa) remains unknown. METHODS: Expansion, activation, and cryopreservation of allogeneic human NK cells obtained from peripheral blood were performed as we previously described. In vitro cytotoxicity was evaluated using the cell counting kit-8. The levels of perforin, granzyme B, interferon-γ, tumor necrosis factor-α, and chemokines (C-C-motif ligand [CCL]1, CCL2, CCL20, CCL3L1, and CCL4; C-X-C-motif ligand [CXCL]1, CXCL16, CXCL2, CXCL3, and CXCL8; and X-motif ligand 1 and 2) were determined using enzyme-linked immunosorbent assay. The expression of CD107a, major histocompatibility complex class I (MHC-I), MHC-I polypeptide-related sequences A and B (MICA/B), cytomegalovirus UL16-binding protein-2/5/6 (ULBP-2/5/6), B7-H6, CD56, CD69, CD25, killer cell Ig-like receptors (KIR)2DL1, KIRD3DL1, NKG2D, NKp30, NKp46, and CD16 of NK cells or BCa and normal urothelial cells were detected using flow cytometry. Cytotoxicity was evaluated using lactate dehydrogenase assay in patient-derived organoid models. BCa growth was monitored in vivo using calipers in male NOD-scid IL2rg-/- mice subcutaneously injected with 5637 and NK cells. Differential gene expressions were investigated using RNA sequence analysis. The chemotaxis of T cells was evaluated using transwell migration assays. RESULTS: We revealed that the NK cells possess higher cytotoxicity against BCa lines with more production of cytokines than normal urothelial cells counterparts in vitro, demonstrated by upregulation of degranulation marker CD107a and increased interferon-γ secretion, by MICA/B/NKG2D and B7H6/NKp30-mediated activation. Furthermore, NK cells demonstrated antitumor effects against BCa in patient-derived organoids and BCa xenograft mouse models. NK cells secreted chemokines, including CCL1/2/20, to induce T-cell chemotaxis when encountering BCa cells. CONCLUSIONS: The expanded NK cells exhibit potent cytotoxicity against BCa cells, with few toxic side effects on normal urothelial cells. In addition, NK cells recruit T cells by secreting a panel of chemokines, which supports the translational application of NK cell intravesical instillation after TURBT from bench to bedside for NMIBC treatment.

FGF21 prevents neuronal cell ferroptosis after spinal cord injury by activating the FGFR1/ß-Klotho pathway.

Spinal cord injury (SCI) is a kind of traumatic nervous system disease caused by neuronal death, causing symptoms like sensory, motor, and autonomic nerve dysfunction. The recovery of neurological function has always been a intractable problem that has greatly distressed individuals and society. Although the involvement of iron-dependent lipid peroxidation leading to nerve cell ferroptosis in SCI progression has been reported, the underlying mechanisms remain unaddressed. Thus, this study aimed to investigate the potential of recombinant human FGF21 (rhFGF21) in inhibiting ferroptosis of nerve cells and improving limb function after SCI, along with its underlying mechanisms. In vivo animal model showed that FGFR1, p-FGFR1, and ß-Klotho protein gradually increased over time after injury, reaching a peak on the third day. Moreover, rhFGF21 treatment significantly reduced ACSL4, increased GPX4 expression, reduced iron deposition, and inhibited ferroptosis. Meanwhile, rhFGF21 decreased cell apoptosis following acute spinal cord damage. In contrast, FGFR1 inhibitor PD173074 partially reversed the rhFGF21-induced therapeutic effects. Overall, this work revealed that rhFGF21 activates the FGFR1/ß-Klotho pathway to decrease ferroptosis of nerve cells, suggesting that FGF21 could be a new therapeutic target for SCI neurological rehabilitation.

TIGIT immune checkpoint blockade enhances immunity of human peripheral blood NK cells against castration-resistant prostate cancer.

Castration-resistant prostate cancer (CRPC) patients have a 14-month median survival, emphasizing the need for alternative treatments. Previously, we demonstrated that expanded high-dose natural killer (NK) cells derived from human peripheral blood exhibit therapeutic efficacy against CRPC. However, which immune checkpoint blockade promotes NK cell antitumor immunity against CRPC remains unknown. Here, we explored immune checkpoint molecule expression in NK and CRPC cells during their interactions, and identified that the T cell immunoreceptor with immunoglobulin and immunoreceptor tyrosine-based inhibition motif domain (TIGIT) monoclonal antibody (mAb), vibostolimab, significantly enhanced NK cell cytotoxicity against CRPC cells and cytokine production in vitro, demonstrated by upregulation of degranulation marker CD107a and Fas-ligand (Fas-L) and increased interferon-gamma (IFN-γ) and tumor necrosis factor-alpha secretion. TIGIT blockade increased Fas-L expression and IFN-γ production via the NF-κB signaling pathway and restored degranulation via the mitogen-activated protein kinase ERK (extracellular signal-regulated kinase) kinase/ERK pathway in activated NK cells. Vibostolimab significantly enhanced NK cell antitumor effects against CRPC in two xenograft mouse models. Vibostolimab also increased T cell chemotaxis induced by activated NK cells in vitro and in vivo. Overall, blocking TIGIT/CD155 signaling enhances the antitumor effect of expanded NK cells against CRPC; this finding supports the translational application of TIGIT mAb and NK cell combination strategies from bench to bedside for CRPC treatment.

Inhibition of Connexin43 Improves the Recovery of Spinal Cord Injury Against Ferroptosis via the SLC7A11/GPX4 Pathway.

Ferroptosis plays a key role in the process of spinal cord injury (SCI). As a signal amplifier, connexin 43 (CX43) participates in cell death signal transduction and aggravates the propagation of injury. However, it remains unclear whether CX43 plays a regulatory role in ferroptosis after SCI. The SCI rat model was established by an Infinite Vertical Impactor to investigate the role of CX43 in SCI-induced ferroptosis. Ferrostatin-1 (Fer-1), an inhibitor of ferroptosis, and a CX43-specific inhibitor (Gap27) were administered by intraperitoneal injection. Behavioral analysis was assessed according to the Basso-Beattie-Bresnahan (BBB) Motor Rating Scale and the inclined plate test. The levels of ferroptosis-related proteins were estimated by qRT-PCR and western blotting, while the histopathology of neuronal injury induced by SCI was evaluated by immunofluorescence, Nissl, FJB and Perl's Blue staining. Meanwhile, transmission electron microscopy was used to observe the ultrastructural changes characteristic of ferroptosis. Gap27 strongly inhibited ferroptosis and therefore improved the functional recovery of SCI, which was similar to the treatment of Fer-1. Notably, the inhibition of CX43 decreased P-mTOR/mTOR expression and reversed the decrease in SLC7A11 induced by SCI. As a result, the levels of GPX4 and glutathione (GSH) increased, while the levels of the lipid peroxidation products 4-hydroxynonenal (4-HNE) and malondialdehyde (MDA) decreased. Together, inhibition of CX43 could alleviate ferroptosis after SCI. These findings reveal a potential mechanism of the neuroprotective role of CX43 after SCI and provide a new theoretical basis for clinical transformation and application.

Association of serum fibroblast growth factor 19 levels with arteriosclerosis parameters assessed by arterial stiffness and atherogenic index of plasma in patients with type 2 diabetes.

BACKGROUND: The role of serum fibroblast growth factor 19 (FGF19) in arteriosclerosis is not well known. In the present study, we aimed to explore whether serum FGF19 levels were related to arteriosclerosis parameters, including arterial stiffness and atherogenic index of plasma (AIP), in patients with type 2 diabetes (T2D). METHODS: A total of 200 patients with type 2 diabetes and 50 healthy controls were recruited for this study from Apr 2017 to Oct 2018. Serum FGF19 levels, arterial stiffness assessed by brachial ankle pulse wave velocity (baPWV), and AIP assessed by the triglyceride to high-density lipoprotein cholesterol (TG/HDL-c) ratio were measured in those subjects. In addition, other relevant clinical data were also collected. RESULTS: Serum FGF19 levels in T2D patients were significantly lower than those in healthy controls (p < 0.05). The arteriosclerosis parameters, including baPWV and AIP, significantly decreased across ascending tertiles of serum FGF19 levels (all p for trend < 0.001). Moreover, the baPWV and AIP were all inversely correlated with serum FGF19 levels (r = - 0.351 and - 0.303, respectively, p < 0.001). Furthermore, after adjusting for other clinical covariates by multiple linear regression analyses, the serum FGF19 levels were independently associated with baPWV (ß = - 0.20, t = - 2.23, p = 0.029) and AIP (ß = - 0.28, t = - 2.66, p = 0.010). CONCLUSIONS: The serum FGF19 levels were independently and inversely associated with baPWV and AIP, which indicate that serum FGF19 may have a protective role in atherosclerosis in patients with T2D.

Polysaccharide from Echinacea purpurea reduce the oxidant stress in vitro and in vivo.

Echinacea purpurea polysaccharides (EPPS) were extracted through water extract and alcohol precipitate method. Three polysaccharides were purified by DEAE cellulose, named EPPS-1, EPPS-2 and EPPS-3. The antioxidant activities in three polysaccharides were screened by free radical scavenging test and EPPS-3 possessed the best antioxidant function. Then the antioxidant activities of EPPS-3 were further explored in oxidative damage model in vitro and in vivo for the first time. The results showed that the antioxidases and the metabolism content were significantly improved in EPPS-3 group. EPPS-3 could protect hepatic tissue from the injury of CCl4 compared with the oxidative damage model. The mechanism research demonstrated that EPPS-3 restrained cell apoptosis and promoted Nrf2 cell signal pathway to play an antioxidant impact. Therefore, EPPS-3 an ingredient could be served as amazing gift for food industry and feed additive.

Serum fibroblast growth factor 19 and endogenous islet beta cell function in type 2 diabetic patients.

BACKGROUND: Fibroblast growth factor 19 (FGF19) takes part in maintaining the balance of glycolipids and may be involved in regulating the secretory activity of islet beta cells in patients with type 2 diabetes. This study aimed to evaluate the relationship between the levels of serum FGF19 and endogenous islet beta cell function in type 2 diabetic patients. METHODS: Samples were obtained from 271 subjects: 85 drug-naïve type 2 diabetes participants exclusively on lifestyle intervention (N-DM group), 122 type 2 diabetes subjects previously used medications (DM group) and 64 normal controls (NC group). Serum FGF19 concentrations were measured by ELISA. The insulin sensitivity (MI), insulin secretion (AUCins/AUCglu) and insulin secretion-sensitivity index-2 (ISSI-2) were also measured in the N-DM and DM. RESULTS: Serum FGF19 levels decreased, in order, from the NC group [median (interquartile range), 245.03 (126.23-317.43) pg/mL] to the N-DM group [170.05 (89.01-244.70) pg/mL] and, finally, to the DM group [142.25 (55.55-187.58) pg/mL] (p for trend < 0.05). Among subjects in the DM group, there was a positive trend in the serum FGF19 concentration; plasma insulin levels at 60 min, 120 min (INS60, INS120, respectively); and area under the insulin curve (AUCins) at two points (r = 0.214, p = 0.025; r = 0.189, p = 0.048; r = 0.188, p = 0.049). However, the differences were no longer observed among the N-DM subjects. Simultaneously, the ISSI-2 was closely related to the serum FGF19 levels (r = 0.297, p = 0.002) among DM subjects. Furthermore, after adjusting for age, sex, duration, therapy and other clinical factors via multiple logistic regression analysis, ISSI-2 was a key independent factor in the levels of FGF19 (ß = 0.281, t = 2.557, p = 0.013). CONCLUSIONS: The serum FGF19 level has a close relation with endogenous beta cell function among DM subjects, as assessed by the ISSI-2. As ISSI-2 is higher in N-DM group, FGF19 may be a main protector in dysfunction of beta cell.

A20 protects neuronal apoptosis stimulated by lipopolysaccharide-induced microglial exosomes.

LPS-induced microglial activation has a major influence on neuronal damage in the inflammatory process. Integral to this is the cellular and molecular interaction between microglia and neurons. Exosomes, a mediator of communication between cells, can transfer lipids, proteins and nucleic acids, affecting many donor and recipient cells. To investigate the mechanism by which microglial exosomes regulate neuronal inflammation after traumatic brain injury, this study primarily analyzed the effect of microglial exosomes on neuronal apoptosis. Exosomes derived from lipopolysaccharide (LPS)-activated microglial cultures were identified and purified. Neurons treated with these exosomes underwent apoptosis. A20 (also known as TNF-inducible protein 3, TNFAIP3) is a deubiquitinating enzyme with key anti-inflammatory functions. A20 is of huge significance to the degeneration and development of neuron. Importantly, A20 protects the exosomes-induced neuronal death, while A20 knockdown increases neuronal death. This study shows that exosomes may be critical for communication between microglia and neurons.

Kaempferol-3-O-glucorhamnoside inhibits inflammatory responses via MAPK and NF-κB pathways in vitro and in vivo.

Klebsiella pneumoniae causes severe infections including pneumonia and sepsis and treatments are complicated by increased levels of antibiotic resistance. We have identified a flavonoid kaempferol-3-O-glucorhamnoside derived from the plant Thesium chinense Turcz that possessed potent anti-inflammatory effects in K. pneumoniae infected mice. Administration of kaempferol-3-O-glucorhamnoside before bacterial challenge effectively suppressed expression of the major inflammatory cytokines TNF-α, IL-6, IL-1ß and PGE2 and ameliorated lung edema. In addition, administration of this compound to cultured RAW macrophages or Balb/c mice resulted in the suppression of NFκB and MAP kinase phosphorylation indicating an inhibitory effect on inflammation in vitro and in vivo. Kaempferol-3-O-glucorhamnoside also decreased ROS levels and overall oxidative stress in lungs and in cultured cells generated by K. pneumoniae exposure. Taken together, kaempferol-3-O-glucorhamnoside is a potent anti-inflammatory in vitro and in vivo and is a promising therapeutic agent for treating K. pneumoniae infections in the clinic.

A Comparison of Two Methods for the Preparation Cefquinome-Loaded Gelatin Microspheres for Lung Targeting.

PURPOSE: The aim of this study was to prepare CEQ-loaded gelatin microspheres and compare two preparation methods, evaluate targeting to the lungs. METHODS: Gelatin microspheres containing CEQ were prepared by an emulsion cross-linking method (ECLM) and a spray-drying method (SDM) and were characterized in terms of morphology, size, drug-loading coefficient, encapsulation ratio and in vitro release. RESULTS: The microspheres prepared by ECLM gave a drug loading (DL) of 19.4 ± 2.4% and an entrapment efficiency (EE) of 80.8 ± 3.2%. The microspheres prepared by SDM resulted in a DL value of 20.8 ± 2.7% and an EE of 95.3 ± 3.8%. The average particle size of microspheres was 7-30 µm by both methods and both preparations sustained CEQ release for 36 h in the target tissue (lungs). The in vitro release profile of the microspheres matched the Korsmeyer-Peppas release pattern. In vivo studies identified the lung as the target tissue and the region of maximum CEQ release. Histopathological examination showed a partial lung inflammation that disappeared spontaneously as the microspheres were biodegraded. In general, the formulations were safe. CONCLUSION: The well-sustained CEQ release from the microspheres revealed its suitability as a drug delivery vehicle that minimized injury to healthy tissues while achieving the accumulation of therapeutic drug for lung targeting. The intravenous administration of CEQ gelatin microspheres prepared by SDM is of potential value in treating lung diseases in animals.