Extracellular vesicles have received extensive attention in the field of bone defect regeneration and repair in recent years. However, natural extracellular vesicles have deficiencies in sustained controlled release, tissue targeting, and drug loading capacity. Therefore, the introduction of engineering strategies to modify extracellular vesicles to enhance their therapeutic efficacy has become a research hotspot.

To review the role and application progress of engineered extracellular vesicles in the regeneration and repair of bone defects.

PubMed, Web of Science, CNKI, and WanFang databases were searched for relevant articles published in the past fifteen years. The search terms were “engineering, extracellular vesicles, exosomes, bone defect, bone regeneration, bone repair” in Chinese and English. After removal of poorly related, outdated, and duplicate studies by screening, 93 articles were finally included for review according to inclusion criteria.

(1) Extracellular vesicles are primarily isolated based on their density, size, immunoaffinity, and surface charge. After isolation, extracellular vesicles are characterized using imaging techniques, size- and counting-based techniques, and flow cytometry. (2) Extracellular vesicles stimulate bone regeneration by regulating immunity, angiogenesis, and proliferation and differentiation of target cells. (3) The engineering strategies of extracellular vesicles include surface modification and cargo loading. (4) The introduction of bone morphogenetic protein 2, mutant hypoxia-inducible factor-1α, vascular endothelial growth factor, miRNA and other bioactive factors into extracellular vesicles through engineering strategies can enhance their regenerative repair ability for bone defects.

Osteoarthritis is a progressive joint condition identified by ongoing deterioration of the cartilage matrix, and there is currently no effective drug treatment plan. Metformin-modified exosomes isolated from bone marrow-derived mesenchymal stem cells can become a new method for treating osteoarthritis due to their avoidance of oral drug adverse reactions and immunogenicity.

To study the controlling impact of exosomes from metformin-altered bone marrow-derived mesenchymal stem cells on chondrocytes.

Rabbit bone marrow-derived mesenchymal stem cells and chondrocytes were cultured in vitro. Bone marrow-derived mesenchymal stem cells derived exosomes and metformin pretreated bone marrow-derived mesenchymal stem cells derived exosomes were collected using a high-speed centrifuge. Chondrocytes were cultured with exosome-containing culture medium for 24 hours and then treated with 100 µmol/L H₂O₂ for 24 hours. The capability changes of two extracellular vesicles on chondrocyte proliferation and migration were detected using CCK8 assay and scratch healing experiment, respectively. Western blot analysis and RT-qPCR were employed to examine the alterations in the expression of type II collagen, P16 protein, and their mRNA in chondrocytes.

Western blot analysis was utilized to assess the changes in the expression of MKK7/JNK pathway proteins. ELISA kits were utilized to measure the activity of cell superoxide dismutase and the levels of malondialdehyde in chondrocytes.

(1) In an oxidative stress environment, the proliferation and migration abilities of chondrocytes were weakened. The two types of exosomes could restore the proliferation and migration abilities of chondrocytes to a certain extent. Metformin pretreated bone marrow-derived mesenchymal stem cells derived exosomes had a significantly better improvement effect (P < 0.05). (2) Compared with normal bone marrow mesenchymal stem cell-derived exosomes, metformin pretreated bone marrow-derived mesenchymal stem cells derived exosomes could more effectively increase type II collagen expression and superoxide dismutase activity (P < 0.05), and were also more effective in reducing P16 expression and malondialdehyde levels (P < 0.05). (3) The two types of exosomes could inhibit the expression of MKK7 and p-JNK proteins to a certain extent, and the inhibitory effect of metformin pretreated bone marrow-derived mesenchymal stem cells derived exosomes was more significant (P < 0.05). The results show that in an oxidative stress environment, metformin pretreated bone marrow-derived mesenchymal stem cells derived exosomes resist chondrocyte aging and promote chondrocyte proliferation by inhibiting the MKK7/JNK pathway.

Erythropoietin/erythropoietin receptor signaling pathway not only participates in bone marrow hematopoiesis, but also regulates the metabolic response of non-hematopoietic tissues, such as brain, heart, skeletal muscle, and adipose tissue. Simultaneously, it can accelerate the mineralization process of periodontal ligament stem cells and reduce oxidative stress damage. However, the mechanism of action on osteogenic differentiation of periodontal ligament stem cells is still unclear.

To investigate the effect and action mechanism of erythropoietin/erythropoietin receptor signaling pathway on osteogenic differentiation of periodontal ligament stem cells.

Enzyme digestion method was used to isolate and culture periodontal ligament stem cells from periodontal disease patients and healthy people. The mRNA and protein levels of erythropoietin receptor in two kinds of periodontal ligament stem cells were detected by qRT-PCR and western blot assay. Erythropoietin receptor expression was silenced by small interfering RNA (siRNA) or activated by erythropoietin. qRT-PCR and western blot assay were used to detect the expression of erythropoietin receptor, levels of osteogenic marker genes Runt-related transcription factor 2 (Runx2), osteocalcin, osteopontin, and bone sialoprotein. Alkaline phosphatase staining and alizarin red staining were applied to measure osteogenic differentiation ability of periodontal ligament stem cells. The phosphorylation of signal transducer and activator of transcription 5 (STAT5) was detected by western blot assay.

(1) The results of qRT-PCR and western blot assay showed that the mRNA and protein levels of erythropoietin receptor in periodontal ligament stem cells in the disease group were significantly lower than those in periodontal ligament stem cells in the healthy group. (2) Alkaline phosphatase staining and alizarin red staining showed that knocking down the erythropoietin receptor can inhibit the osteogenic differentiation ability of periodontal ligament stem cells. qRT-PCR results showed that compared with the control group, knockdown of the erythropoietin receptor group significantly reduced expression levels of Runt-related transcription factor 2, osteocalcin, osteopontin, and bone sialoprotein (P < 0.05). (3) qRT-PCR results showed that after erythropoietin treatment, the expression of erythropoietin receptor in periodontal ligament stem cells recovered. Silencing erythropoietin receptor and then administration of erythropoietin treatment reversed the expression level of erythropoietin receptor. Erythropoietin treatment increased the osteogenic differentiation ability of periodontal ligament stem cells in the disease group and the expression level of the osteogenic marker gene Runt-related transcription factor 2 (P < 0.05). Silencing the expression of STAT5 inhibited this effect of erythropoietin. (4) Western blot assay results showed that with the extension of erythropoietin treatment time, the phosphorylation level of STAT5 increased in periodontal ligament stem cells in the disease group (P < 0.05). The above results indicate that erythropoietin restores the osteogenic differentiation ability of pathological periodontal ligament stem cells by inducing the phosphorylation of STAT5.

Previous studies have shown that Huosui Formula has a synergistic effect on the immune and hematopoietic regulation of patients with myelodysplastic syndrome, but the specific mechanism is not yet clear.

To explore the effect and mechanism of Huosui Formula on bone marrow hematopoiesis in rats with myelodysplastic syndrome.

A total of 70 SD rats were randomly divided into a normal control group (n=10), a model group (n=15), a western medicine group (n=15), a low-dose Huosui Formula group (n=15), and a high-dose Huosui Formula group (n=15). Except for the normal control group, the other four groups were injected with dimethylbenzanthracene via the tail vein to induce the establishment of rat myelodysplastic syndrome models. After modeling, the normal control group and the model group were given normal saline; the western medicine group was given thalidomide capsules 10 mg/kg and retinoic acid tablets 4 mg/kg, and the low-dose Huosui Formula group and the high-dose Huosui Formula group were given 1.5 and 6 g/kg Huosui Formula, respectively, by intragastric administration once a day for 28 consecutive days. Peripheral blood and femoral bone marrow tissue were collected to detect peripheral blood routine and bone marrow biopsy hematopoietic proliferation. Flow cytometry was used to detect T lymphocyte subsets and the expression of CTLA-4 and PD-1 on T lymphocytes.

(1) Compared with the normal control group, peripheral blood leukocyte, neutrophil, hemoglobin, platelet, and CD4⁺, CD4⁺/CD8⁺ levels were decreased in the model group significantly (P < 0.05), while CD4⁺PD-1⁺, CD8⁺PD-1⁺, CD4⁺CTLA-4⁺, and CD8⁺CTLA-4⁺ expressions were significantly upregulated (P < 0.05). (2) In all dosage groups, myelopoietic proliferation was increased compared with the model group, with no significant difference between the groups (P > 0.05). (3) Compared with the model group, leukocytes, hemoglobin, platelets, and CD4⁺, CD4⁺/CD8⁺ were significantly elevated in the high-dose Huosui Formula group (P < 0.05), the expression of CD8⁺ was significantly lower (P < 0.05), and the levels of CD4⁺PD-1⁺, CD8⁺PD-1⁺, CD4⁺CTLA-4⁺, and CD8⁺CTLA-4⁺ were down-regulated but not statistically significant (P > 0.05). (4) The western medicine group and the high-dose Huosui Formula group showed similar efficacy. The improvement of each index in the high-dose Huosui Formula group was superior to that in the low-dose Huosui Formula group. These findings indicate that Huosui Formula can improve the bone marrow hematopoiesis in myelodysplastic syndrome model rats, increase the levels of CD4⁺, and CD4⁺/CD8⁺ while down-regulate the expression levels of CD4⁺PD-1⁺, CD8⁺PD-1⁺, CD4⁺CTLA-4⁺, and CD8⁺CTLA-4⁺. These observations suggest a link to the negative immunoregulation mechanism.

Numerous studies have indicated that pyroptosis plays a key role in the progression of cancer. In recent years, research has shown that pyroptosis is inextricably linked to the occurrence, development, and treatment of breast cancer. The development of effective pyroptosis-based therapeutic strategies has become a hot topic in the field of breast cancer treatment.

To comprehensively analyze the mechanisms of pyroptosis, explore the role of pyroptosis in the anti-tumor effects in breast cancer, and its potential application value in clinical treatment.

Using English search terms “pyroptosis, breast cancer, inflammasome, gasdermin, caspase, drug resistance, treatment”, PubMed database was searched for articles published from inception to August 2024. Through the preliminary screening of reading titles and abstracts, literature with poor relevance to the research content, outdated information, repeated views, and lack of authority was excluded. Finally, 121 articles were included for review.

Pyroptosis is a special form of programmed cell death that is carried out by the activation of the gasdermin family of proteins, showing potential application value in the treatment of breast cancer. Long-term or improper treatment can lead to drug resistance in cancer cells; research on the mechanism of pyroptosis helps to overcome resistance deficiencies. Pyroptosis can trigger immunogenic cell death, promoting the release of tumor-specific antigens, thereby activating the immune system and enhancing its ability to recognize and clear tumor cells. The expression levels of pyroptosis-related genes can serve as prognostic indicators for breast cancer, helping to assess patients’ treatment responses and survival periods. Research on the mechanisms of pyroptosis can provide new strategies for the treatment of breast cancer, such as targeted drugs and therapeutic methods that induce pyroptosis, contributing to the realization of personalized treatment plans for breast cancer.