Biomechanical Characteristics and Analysis Approaches of Bone and Bone Substitute Materials.

Bone has a special structure that is both stiff and elastic, and the composition of bone confers it with an exceptional mechanical property. However, bone substitute materials that are made of the same hydroxyapatite (HA) and collagen do not offer the same mechanical properties. It is important for bionic bone preparation to understand the structure of bone and the mineralization process and factors. In this paper, the research on the mineralization of collagen is reviewed in terms of the mechanical properties in recent years. Firstly, the structure and mechanical properties of bone are analyzed, and the differences of bone in different parts are described. Then, different scaffolds for bone repair are suggested considering bone repair sites. Mineralized collagen seems to be a better option for new composite scaffolds. Last, the paper introduces the most common method to prepare mineralized collagen and summarizes the factors influencing collagen mineralization and methods to analyze its mechanical properties. In conclusion, mineralized collagen is thought to be an ideal bone substitute material because it promotes faster development. Among the factors that promote collagen mineralization, more attention should be given to the mechanical loading factors of bone.

Long Noncoding RNA MIR4435-2HG Suppresses Colorectal Cancer Initiation and Progression By Reprogramming Neutrophils.

MIR4435-2HG, also known as LINC00978, has previously been described as an oncogenic long noncoding RNA (lncRNA). However, we show here that Mir4435-2hg depletion promoted colorectal tumorigenesis and progression in in vivo models of colitis-associated colorectal cancer, spontaneous intestinal adenomatous polyposis, and subcutaneous tumors. Alteration of MIR4435-2HG in colorectal cancer cells did not change the potential for cell proliferation, migration, or invasion in vitro. RNAscope assays showed that most MIR4435-2HG was located in the tumor stroma, which caused high expression of MIR4435-2HG in colorectal cancer tumor tissue. Transcriptome analysis of colorectal cancer tissues from wild-type and Mir4435-2hg-deficient mice revealed Mir4435-2hg as a tumor suppressor gene that regulated the immune microenvironment. Loss of Mir4435-2hg led to a decline in neutrophils and elevation of polymorphonuclear myeloid-derived suppressor cells (PMN-MDSC). In tissue-specific Mir4435-2hg knockout mice, we confirmed that Mir4435-2hg depletion in neutrophils, but not in intestinal epithelial cells, promoted colorectal cancer progression. Mechanistically, Mir4435-2hg depletion enhanced the immunosuppressive ability of PMN-MDSCs by disturbing their fatty acid metabolism. These findings suggest that MIR4435-2HG is a tumor-suppressing lncRNA whose deficiency could increase tumor-infiltrating PMN-MDSCs and enhance the immunosuppressive potential of PMN-MDSCs to promote colorectal cancer development. This provides a theoretical basis for further illustrating the pathogenesis of colorectal cancer and a potential antitumor immunotherapy target.

Physical and Chemical Characterization of Biomineralized Collagen with Different Microstructures.

Mineralized collagen is the basic unit in hierarchically organized natural bone with different structures. Polyacrylic acid (PAA) and periodic fluid shear stress (FSS) are the most common chemical and physical means to induce intrafibrillar mineralization. In the present study, non-mineralized collagen, extrafibrillar mineralized (EM) collagen, intrafibrillar mineralized (IM) collagen, and hierarchical intrafibrillar mineralized (HIM) collagen induced by PAA and FSS were prepared, respectively. The physical and chemical properties of these mineralized collagens with different microstructures were systematically investigated afterwards. Transmission electron microscopy (TEM) and scanning electron microscopy (SEM) showed that mineralized collagen with different microstructures was prepared successfully. The pore density of the mineralized collagen scaffold is higher under the action of periodic FSS. Fourier transform infrared spectroscopy (FTIR) analysis showed the formation of the hydroxyapatite (HA) crystal. A significant improvement in the pore density, hydrophilicity, enzymatic stability, and thermal stability of the mineralized collagen indicated that the IM collagen under the action of periodic FSS was beneficial for maintaining collagen activity. HIM collagen fibers, which are prepared under the co-action of periodic FSS and sodium tripolyphosphate (TPP), may pave the way for new bone substitute material applications.

Orthophosphate and alkaline phosphatase induced the formation of apatite with different multilayered structures and mineralization balance.

Mineralized collagen is a natural organic-inorganic composite. The combination of organic collagen and inorganic apatite to form different nanostructures is the key to producing bone substitutes with biomechanical properties that are as identical to normal bone as possible. However, the formation of apatite with different nanostructures during collagen mineralization is unexplored. Here, pyrophosphate (Pyro-P), as an important hydrolysate of adenosine triphosphate in the body, was introduced to prepare mineralized collagen under the regulation of alkaline phosphatase (ALP) and orthophosphate (Ortho-P). Scanning electron microscopy (SEM) and transmission electron microscopy (TEM) results showed that mineralized collagen, which combined with different crystallinities and multilayered structured apatite, was successfully prepared. A combination of ion chromatography (IC), Fourier transform infrared (FTIR) spectroscopy, circular dichroism (CD), and thermogravimetry (TG) analyses revealed the crucial role of Ortho-P in the formation of multilayered flower-shaped apatite with different crystallinities and in the maintenance of mineralization balance. Mineralization balance is of great significance for maintaining normal bone morphology during bone regeneration. Overall, our results provide a promising method to produce new bone substitute materials for the repair of large bone defects and a deeper insight into the mechanisms of biomineralization.

Inhibition of RhoA/Rho kinase signaling pathway by fasudil protects against kainic acid-induced neurite injury.

AIM: RhoA/Rho kinase pathway is essential for regulating cytoskeletal structure. Although its effect on normal neurite outgrowth has been demonstrated, the role of this pathway in seizure-induced neurite injury has not been revealed. The research examined the phosphorylation level of RhoA/Rho kinase signaling pathway and to clarify the effect of fasudil on RhoA/Rho kinase signaling pathway and neurite outgrowth in kainic acid (KA)-treated Neuro-2A cells and hippocampal neurons. METHOD: Western blotting analysis was used to investigate the expression of key proteins of RhoA/Rho kinase signaling pathway and the depolymerization of actin. After incubated without serum to induce neurite outgrowth, Neuro-2A cells were fixed, and immunofluorescent assay of rhodamine-phalloidin was applied to detect the cellular morphology and neurite length. The influence of KA on neurons was detected in primary hippocampal neurons. Whole-cell patch clamp was conducted in cultured neurons or hippocampal slices to record action potentials. RESULT: KA at the dose of 100-200 µmol/L induced the increase in phosphorylation of Rho-associated coiled-coil-containing protein kinase and decrease in phosphorylation of Lin11, Isl-1 and Mec-3 kinase and cofilin. The effect of 200 µmol/L KA was peaked at 1-2 hours, and then gradually returned to baseline after 8 hours. Pretreatment with Rho kinase inhibitor fasudil reversed KA-induced activation of RhoA/Rho kinase pathway and increase in phosphorylation of slingshot and 14-3-3, which consequently reduced the ratio of G/F-actin. KA treatment induced inhibition of neurite outgrowth and decrease in spines both in Neuro-2a cells and in cultured hippocampal neurons, and pretreatment with fasudil alleviated KA-induced neurite outgrowth inhibition and spine loss. CONCLUSION: These data indicate that inhibiting RhoA/Rho kinase pathway might be a potential treatment for seizure-induced injury.

Crucial roles of graphene oxide in preparing alginate/nanofibrillated cellulose double network composites hydrogels.

In this study, a novel strategy to prepare sodium alginate (SA)/nano fibrillated cellulose (NFC) double network (DN) hydrogel beads with the aid of graphene oxide (GO) was developed. In comparison with the multi-step freezing-thawing method, this study employs a facile one-step freeze drying method with the presence of GO sheets. The crucial roles of GO were highlighted as an efficient nucleating agent of NFC and a reinforcer for the hydrogel. The adsorption property of the DN hydrogel towards crystal violet (CV) was also studied. Results indicated that the introduction of GO could greatly facilitate the formation of double networks. Furthermore, the as-prepared DN hydrogel beads exhibited an efficacious adsorption property towards CV. The maximum adsorption capacity of the hydrogels for CV was observed as 665 mg g-1. Therefore, our approach here represents a facile method for the preparation of crystalline polymer based DN hydrogels to replace the awkward freezing-thawing process, giving inspiration for DN hydrogels design and preparation. Moreover, due to its efficient adsorption capacity, the hydrogels hold great promise for the water pollution control materials.

The inflammatory cytokine IL-6 induces FRA1 deacetylation promoting colorectal cancer stem-like properties.

Colorectal cancer (CRC) has long been known for its tight association with chronic inflammation, thought to play a key role in tumor onset and malignant progression through the modulation of cancer stemness. However, the underlying molecular and cellular mechanisms are still largely elusive. Here we show that the IL-6/STAT3 inflammatory signaling axis induces the deacetylation of FRA1 at the Lys-116 residue located within its DNA-binding domain. The HDAC6 deacetylase underlies this key modification leading to the increase of FRA1 transcriptional activity, the subsequent transactivation of NANOG expression, and the acquisition of stem-like cellular features. As validated in a large (n = 123) CRC cohort, IL-6 secretion was invariably accompanied by increased FRA1 deacetylation at K116 and an overall increase in its protein levels, coincident with malignant progression and poor prognosis. Of note, combined treatment with the conventional cytotoxic drug 5-FU together with Tubastatin A, a HDAC6-specific inhibitor, resulted in a significant in vivo synergistic inhibitory effect on tumor growth through suppression of CRC stemness. Our results reveal a novel transcriptional and posttranslational regulatory cross-talk between inflammation and stemness signaling pathways that underlie self-renewal and maintenance of CRC stem cells and promote their malignant behavior. Combinatorial treatment aimed at the core regulatory mechanisms downstream of IL-6 may offer a novel promising approach for CRC treatment.

[Rapamycin treatment starting at 24 h after cerebral ischemia/reperfusion exhibits protective effect on brain injury in rats].

OBJECTIVE: To investigate whether rapamycin treatment starting at 24 h after cerebral ischemia/reperfusion(I/R) has protective effect on brain injury in rats. METHODS: The rat I/R model was established by middle cerebral artery occlusion according to Longa's method. A total of 104 Sprague Dawley rats were randomly divided into sham group, model group, and rapamycin-treated groups (6 h or 24 h after modeling). Neurological function was assessed with neurological severity score (NSS). Triphenyl tetrazolium chloride (TTC) staining and Fluoro-Jade B (FJB) staining were used to examine the infarct volume and neuronal apoptosis, respectively. The expression of p-S6 protein in mTOR signaling pathway was detected by Western blot analysis. RESULTS: Compared with sham group, NSS of the model group was significantly increased and TTC staining indicated obvious infarct area (all P<0.01). Furthermore, significantly increased number of FJB-positive cells and p-S6 expression in the penumbra area were shown in the model group (all P<0.01). Compared with the model group, both rapamycin-treated groups demonstrated decreased NSS, infarction volume and FJB positive cells as well as p-S6 expression in the penumbra area (P<0.05 or P<0.01). There was no significant difference between the groups of rapamycin administrated 6 h and 24 h after modeling (all P>0.05). CONCLUSIONS: Rapamycin treatment starting at 24 h after I/R exhibits protective effect on brain injury in rats.