Methylated ZNF582 as a triage marker for occult cervical cancer and advanced cervical intraepithelial neoplasia.

Aim: To explore the appropriate triage methods for women infected with high-risk human papillomavirus (hrHPV). Materials & methods: A total of 424 out of 872 hrHPV-infected women were divided into cervicitis (n = 123), cervical intraepithelial neoplasia grade 1 (CIN1; n = 89), CIN2 (n = 72), CIN3 (n = 87) and cervical cancer (n = 53) groups. Results: The sensitivity/specificity of ZNF582m, PAX1m and liquid-based cytology (LBC) for hrHPV-infected women with transformation zone 3 CIN3+ was 83.9/93.1, 77.4/90.6 and 80.6/58.5%, respectively. The ZNF582m/PAX1m test had a higher specificity than LBC (p < 0.001) and similar sensitivity to that observed for LBC (p > 0.05). ZNF582m/PAX1m improved the positive predictive value of CIN3+ (64.7/60.0%) in low-grade LBC (negative predictive value: 91.7/88.7%). Conclusion: ZNF582m was superior to PAX1m and LBC tests in detecting CIN3+ in hrHPV-infected women.

Human papillomavirus (HPV) testing is the main method for cervical cancer screening. Although most HPV infections are transient and can be cleared by the body, persistent infection with HPV can lead to cervical cancer. In this study, 424 HPV-infected women were divided into normal, cervical intraepithelial neoplasia grade 1 (CIN1), CIN2, CIN3 and cervical cancer groups according to the grade of cervical lesion (low to high). Women with CIN3 or cervical cancer need treatment. ZNF582^m, PAX1^m and liquid-based cytology detected 83.9, 77.4 and 80.6% of women with CIN3+ and 93.1, 90.6 and 58.5% of women without CIN3+. However, the ZNF582^m test was superior to the PAX1^m and liquid-based cytology tests.

Management of patients with positive margin after conization for high-grade cervical intraepithelial lesions.

BACKGROUND AND OBJECTIVE: To evaluate the clinical efficacy and safety of 5-aminolevulinic acid photodynamic therapy (ALA-PDT) in the treatment of patients with positive margin in comparison to regular follow-up, and a repeat cervical conization. MATERIALS AND METHODS: A retrospective analysis was conducted using 83 patients with pathologically confirmed high-grade cervical intraepithelial neoplasia (CIN) with a positive margin after conization. The management methods and patient prognosis were analyzed and compared. RESULTS: Thirty-five, 33, and 15 patients were treated for regular follow-up, ALA-PDT, and a repeat cervical conization, respectively. About 33.3% (5/15) patients had residual lesions of low-grade CIN and above after recognization. The clinical characteristics of patients in the three groups were similar. The residual lesion rates of patients selected for follow-up, ALA-PDT, and recognization were 34.3% (12/35), 9.1% (3/33), and 0% (0/15), respectively, at 6-month follow-up (p = 0.004). The HPV clearance rates were 31.3%, 66.7%, and 84.6%, respectively (p = 0.01). Further analysis showed that a positive margin in the inscribed margin of the cervical canal (p = 0.022) and persistent HR-HPV positive tests after initial conization (p = 0.003) significantly increased the risk of residual disease. At 2-year follow-up, the recurrence rates of lesions were 3.3% and 26.1% in the ALA-PDT and follow-up groups, respectively (p = 0.021). Notably, the recurrence rates were not significantly different between the ALA-PDT and recognization groups (3.3% vs. 6.7%) (p = 0.561). CONCLUSION: ALA-PDT is an effective treatment for patients with a positive margin after cervical conization for high-grade CIN. Compared with regular follow-up, ALA-PDT can reduce residual and recurrence rate. Moreover, there was no significant difference in the efficacy between AlA-PDT and recognization.

RNA-Seq Analysis Identifies Differentially Expressed Genes in the Longissimus dorsi of Wagyu and Chinese Red Steppe Cattle.

Meat quality has a close relationship with fat and connective tissue; therefore, screening and identifying functional genes related to lipid metabolism is essential for the production of high-grade beef. The transcriptomes of the Longissimus dorsi muscle in Wagyu and Chinese Red Steppe cattle, breeds with significant differences in meat quality and intramuscular fat deposition, were analyzed using RNA-seq to screen for candidate genes associated with beef quality traits. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis showed that the 388 differentially expressed genes (DEGs) were involved in biological processes such as short-chain fatty acid metabolism, regulation of fatty acid transport and the peroxisome proliferator-activated receptor (PPAR) signaling pathway. In addition, crystallin alpha B (CRYAB), ankyrin repeat domain 2 (ANKRD2), aldehyde dehydrogenase 9 family member A1 (ALDH9A1) and enoyl-CoA hydratase and 3-hydroxyacyl CoA dehydrogenase (EHHADH) were investigated for their effects on intracellular triglyceride and fatty acid content and their regulatory effects on genes in lipogenesis and fatty acid metabolism pathways. This study generated a dataset from transcriptome profiling of two cattle breeds, with differing capacities for fat-deposition in the muscle, and revealed molecular evidence that CRYAB, ANKRD2, ALDH9A1 and EHHADH are related to fat metabolism in bovine fetal fibroblasts (BFFs). The results provide potential functional genes for maker-assisted selection and molecular breeding to improve meat quality traits in beef cattle.

Src-mediated phosphorylation of GAPDH regulates its nuclear localization and cellular response to DNA damage.

Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is a key enzyme involved in energy metabolism. Recently, GAPDH has been suggested to have extraglycolytic functions in DNA repair, but the underlying mechanism for the GAPDH response to DNA damage remains unclear. Here, we demonstrate that the tyrosine kinase Src is activated under DNA damage stress and phosphorylates GAPDH at Tyr41. This phosphorylation of GAPDH is essential for its nuclear translocation and DNA repair function. Blocking the nuclear import of GAPDH by suppressing Src signaling or through a GAPDH Tyr41 mutation impairs its response to DNA damage. Nuclear GAPDH is recruited to DNA lesions and associates with DNA polymerase ß (Pol ß) to function in DNA repair. Nuclear GAPDH promotes Pol ß polymerase activity and increases base excision repair (BER) efficiency. Furthermore, GAPDH knockdown dramatically decreases BER efficiency and sensitizes cells to DNA damaging agents. Importantly, the knockdown of GAPDH in colon cancer SW480 cells and xenograft models effectively enhances their sensitivity to the chemotherapeutic drug 5-FU. In summary, our findings provide mechanistic insight into the new function of GAPDH in DNA repair and suggest a potential therapeutic target in chemotherapy.

IGFBP7 downregulation or overexpression effect on bovine preadipocyte differentiation.

The insulin-like growth factor binding-protein 7 (IGFBP7) has binding affinities to IGFs and is able to either positively or negatively regulate the IGFs signaling pathway. It also plays a crucial role in cell growth, differentiation and development in an IGF-independent manner. Herein, we investigated the specific regulation of the gene encoding for IGFBP7during the differentiation process of the adipocyte cells of the Yan Yellow Cattle by interfering with or by overexpressing the IGFBP7 gene. As a result, we found that the mRNA expression levels of IGFBP7 were significantly increased during the formation of progenitor cells. In addition, the expression levels of the lipoprotein lipase (LPL) and transcription factors (PPARγ, C/EBPα) were also significantly increased. IGFBP7 gene overexpression and RNA interfering promoted and inhibited respectively the lipid accumulation and triglyceride production in mature adipocytes, and the expression of the LPL and transcription factors (PPARγ, C/EBPα). The changes in the protein expression levels of IGFBP7 and adipogenic factors were in accord with the changes observed in the mRNA levels. In conclusion, our results indicate that IGFBP7 plays an important regulatory role in the differentiation of preadipocyte cells.

ALDH1A1 effect on Yan Yellow Cattle preadipocyte differentiation.

Aldehyde dehydrogenase 1A1 (ALDH1A1) is a cytosolic enzyme that mainly catalyzes the oxidation of acetaldehyde into acetic acid and participates in the regulation of differentiation and gene expression in fat cell growth and development. However, the physiological role of ALDH1A1 in the formation of fat cell precursors in the Yan Yellow Cattle is still not clear. Herein, we investigated the specific regulation of the gene encoding for ALDH1A1 during the differentiation process of the adipocyte cells of the Yan Yellow Cattle by interfering or overexpressing the ALDH1A1 gene. As a result, we found that the mRNA expression levels of ALDH1A1 were significantly increased during the formation of progenitor cells. In addition, the expression levels of the Lipoprotein lipase (LPL) and transcription factors (PPARγ, C/EBPα) were also significantly increased. ALDH1A1 gene overexpression and RNA interfering promoted and inhibited respectively the lipid accumulation and triglyceride production in mature adipocytes, and the expression of the LPL and transcription factors (PPARγ, C/EBPα). The changes in the protein expression levels of ALDH1A1 and adipogenic factors were in accord with the changes observed in the mRNA levels. In conclusion, our results indicate that ALDH1A1 plays an important regulatory role in the differentiation of preadipocyte cells.

Humic Acids Limit the Precipitation of Cadmium and Arsenate at the Brushite-Fluid Interface.

Bioavailability and mobility of cadmium (Cd2+) and arsenate (As5+) in soils can be effectively lowered through the dissolution of brushite (dicalcium phosphate dihydrate, CaHPO4·2H2O) coupled with the precipitation of a more stable mineral phase containing both Cd and As. Due to the ubiquitous presence of humic acid (HA) in soil environments, it is more complex to predict the fate of dissolved Cd and As during such sequestration. Here, we used in situ atomic force microscopy (AFM) to image the kinetics of simultaneous precipitation of Cd and As at the brushite-fluid interface in the presence of HA. Results show that HA inhibits the formation of both amorphous and crystalline Cd(5- x)Ca x(PO4)(3- y)(AsO4) y(OH) on the (010) face of brushite. A combination of X-ray photoelectron spectroscopy (XPS) and real-time surface-enhanced Raman spectroscopy (SERS) reveals that part of As5+ reduction into As3+ with HA and [HA-Cd] complexation occurs, modulating the concentrations of free Cd2+ and As5+ ions to inhibit subsequent precipitation of a Cd(5- x)Ca x(PO4)(3- y)(AsO4) y(OH) phase on the dissolving brushite surface. A combination of AFM imaging, SERS analyses, and PhreeqC simulations suggests that environmentally relevant humic substances can limit the precipitation of Cd and As at mineral surfaces through a mechanism of oxidation/reduction and aqueous/surface complexation. This may exacerbate the transportation of these contaminants into waters by subsurface fluid flow, and research attempts to weaken the negative effect of HA are needed.

Dynamics and Molecular Mechanism of Phosphate Binding to a Biomimetic Hexapeptide.

Phosphorus (P) recovery from wastewater is essential for sustainable P management. A biomimetic hexapeptide (SGAGKT) has been demonstrated to bind inorganic P in P-rich environments, however the dynamics and molecular mechanisms of P-binding to the hexapeptide still remain largely unknown. We used dynamic force spectroscopy (DFS) to directly distinguish the P-unbound and P-bound SGAGKT adsorbed to a mica (001) surface by measuring the single-molecule binding free energy (Δ Gb). Using atomic force microscopy (AFM) to determine real-time step retreat velocities of triangular etch pits formed at the nanoscale on the dissolving (010) face of brushite (CaHPO4·2H2O) in the presence of SGAGKT, we observed that SGAGKT peptides promoted in situ dissolution through an enhanced P-binding driven by hydrogen bonds in a P-loop being capable of discriminating phosphate over arsenate, concomitantly forming a thermodynamically favored SGAGKT-HPO42- complexation at pH 8.0 and relatively low ionic strength, consistent with the DFS and isothermal titration calorimetry (ITC) determinations. The findings reveal the thermodynamic and kinetic basis for binding of phosphate to SGAGKT and provide direct evidence for phosphate discrimination in phosphate/arsenate-rich environments.

Direct Observation of Simultaneous Immobilization of Cadmium and Arsenate at the Brushite-Fluid Interface.

Cadmium (Cd2+) and Arsenate (As5+) are the main toxic elements in soil environments and are easily taken up by plants. Unraveling the kinetics of the adsorption and subsequent precipitation/immobilization on mineral surfaces is of considerable importance for predicting the fate of these dissolved species in soils. Here we used in situ atomic force microscopy (AFM) to image the dissolution on the (010) face of brushite (dicalcium phosphate dihydrate, CaHPO4·2H2O) in CdCl2- or Na2HAsO4-bearing solutions over a broad pH and concentration range. During the initial dissolution processes, we observed that Cd or As adsorbed on step edges to modify the morphology of etch pits from the normal triangular shape to a four-sided trapezium. Following extended reaction times, the respective precipitates were formed on brushite through a coupled dissolution-precipitation mechanism. In the presence of both CdCl2 and Na2HAsO4 in reaction solutions at pH 8.0, high-resolution transmission electron microscopy (HRTEM) showed a coexistence of both amorphous and crystalline phases, i.e., a mixed precipitate of amorphous and crystalline Cd(5- x)Ca x(AsO4)(3- y)(PO4) yOH phases was detected. These direct dynamic observations of the transformation of adsorbed species to surface precipitates may improve the mechanistic understanding of the calcium phosphate mineral interface-induced simultaneous immobilization of both Cd and As and subsequent sequestration in diverse soils.

Visualizing Organophosphate Precipitation at the Calcite-Water Interface by in Situ Atomic-Force Microscopy.

Esters of phosphoric acid constitute a large fraction of the total organic phosphorus (OP) in the soil environment and, thus, play an important role in the global phosphorus cycle. These esters, such as glucose-6-phosphate (G6P), exhibit unusual reactivity toward various mineral particles in soils, especially those containing calcite. Many important processes of OP transformation, including adsorption, hydrolysis, and precipitation, occur primarily at mineral-fluid interfaces, which ultimately governs the fate of organophosphates in the environment. However, little is known about the kinetics of specific mineral-surface-induced adsorption and precipitation of organophosphates. Here, by using in situ atomic-force microscopy (AFM) to visualize the dissolution of calcite (1014) faces, we show that the presence of G6P results in morphology changes of etch pits from the typical rhombohedral to a fan-shaped form. This can be explained by a site-selective mechanism of G6P-calcite surface interactions that stabilize the energetically unfavorable (0001) or (0112) faces through step-specific adsorption of G6P. Continuous dissolution at calcite (1014)-water interfaces caused a boundary layer at the calcite-water interface to become supersaturated with respect to a G6P-Ca phase that then drives the nucleation and growth of a G6P-Ca precipitate. Furthermore, after the introduction of the enzyme alkaline phosphatase (AP), the precipitates were observed to contain a mixture of components associated with G6P-Ca, amorphous calcium phosphate (ACP)-hydroxyapatite (HAP) and dicalcium phosphate dihydrate (DCPD). These direct dynamic observations of the transformation of adsorption- and complexation-surface precipitation and enzyme-mediated pathways may improve the mechanistic understanding of the mineral-interface-induced organophosphate sequestration in the soil environment.

Alkaloids of Aconiti Lateralis Radix Praeparata inhibit growth of non-small cell lung cancer by regulating PI3K/Akt-mTOR signaling and glycolysis.

Aconiti Lateralis Radix Praeparata (Fuzi in Chinese) is widely used in the clinical treatment of tumors. This study aims to explore the active fractions and underlying mechanisms of Fuzi in the treatment of non-small cell lung cancer (NSCLC). Fuzi alkaloids (FZA) is prepared and found to inhibit the growth of NSCLC both in vitro and in vivo significantly. A total of 53 alkaloids are identified in FZA by UPLC-Q-TOF-MS. Proteomics experiment show that 238 differentially expressed proteins regulated by FZA are involved in amino acid anabolism, pyrimidine metabolism and PI3K/Akt-mTOR signaling pathway. Metabolomics analyses identify 32 significant differential metabolites which are mainly involved in amino acid metabolism, TCA cycle and other pathways. Multi-omics research combined with molecular biological assays suggest that FZA might regulate glycolysis through PI3K/Akt-mTOR pathway to treat NSCLC. The study lays a foundation for the anti-cancer investigation of Fuzi and provides a possible scientific basis for its clinical application.

Direct imaging of nanoscale dissolution of dicalcium phosphate dihydrate by an organic ligand: concentration matters.

Unraveling the kinetics and mechanisms of sparingly soluble calcium orthophosphate (Ca-P) dissolution in the presence of organic acids at microscopic levels is important for an improved understanding in determining the effectiveness of organic acids present in most rhizosphere environments. Herein, we use in situ atomic force microscopy (AFM) coupled with a fluid reaction cell to image dissolution on the (010) face of brushite, CaHPO4 · 2H2O, in citrate-bearing solutions over a broad concentration range. We directly measure the dependence of molecular step retreat rate on citrate concentration at various pH values and ionic strengths, relevant to soil solution conditions. We find that low concentrations of citrate (10-100 µM) induced a reduction in step retreat rates along both the [100]Cc and [101]Cc directions. However, at higher concentrations (exceeding 0.1 mM), this inhibitory effect was reversed with step retreat speeds increasing rapidly. These results demonstrate that the concentration-dependent modulation of nanoscale Ca-P phase dissolution by citrate may be applied to analyze the controversial role of organic acids in enhancing Ca-P mineral dissolution in a more complex rhizosphere environment. These in situ observations may contribute to resolving the previously unrecognized interactions of root exudates (low molecular weight organic acids) and sparingly soluble Ca-P minerals.

A highlightedly improved method for isolating and characterizing calcium oxalate crystals from tubercles of Mammillaria schumannii.

BACKGROUND: Calcium oxalate (CaOx) is the most prevalent and widespread biomineral in plants and is involved in protective and/or defensive functions against abiotic stress factors. It is, however, expected that this function has an extremely significant contribution to growth processes in plants bearing large amounts of CaOx, such as cacti growing in desert environment. RESULTS: In our research, small-sized CaOx crystals (≤ 20 µm) with tetrahedral or spherical shapes were observed to dominate in each epidermal and cortical cell from the tubercles of Mammillaria schumannii, a species from the Cereoideae subfamily, having tubercles (main photosynthetic organs) united with adjacent ones almost into ridges on its stem. Because they have potential significant functions, differential centrifugations after mechanical blending were used to obtain these small-sized CaOx crystals, which extremely tend to adhere to tissue or suspend in solution. And then the combined Scanning Electron Microscope Energy Dispersive System (SEM-EDS) and Raman spectroscopy were further performed to demonstrate that the extracted crystals were mainly CaC2O4·2H2O. Interestingly, spherical druses had 2 obvious abnormal Raman spectroscopy peaks of -CH and -OH at 2947 and 3290 cm-1, respectively, which may be attributed to the occluded organic matrix. The organic matrix was further extracted from spherical crystals, which could be polysaccharide, flavone, or lipid compounds on the basis of Raman spectroscopy bands at 2650, 2720, 2770, and 2958 cm-1. CONCLUSIONS: Here we used a highlightedly improved method to effectively isolate small-sized CaOx crystals dominating in the epidermal and cortical cells from tubercles of Mammillaria schumannii, which extremely tended to adhere plant tissues or suspend in isolation solution. And then we further clarified the organic matrix getting involved in the formation of CaOx crystals. This improved method for isolating and characterizing biomineral crystals can be helpful to understand how CaOx crystals in cacti function against harsh environments such as strong light, high and cold temperature, and aridity.

The Radiological Physics Center's standard dataset for small field size output factors.

Delivery of accurate intensity-modulated radiation therapy (IMRT) or stereotactic radiotherapy depends on a multitude of steps in the treatment delivery process. These steps range from imaging of the patient to dose calculation to machine delivery of the treatment plan. Within the treatment planning system's (TPS) dose calculation algorithm, various unique small field dosimetry parameters are essential, such as multileaf collimator modeling and field size dependence of the output. One of the largest challenges in this process is determining accurate small field size output factors. The Radiological Physics Center (RPC), as part of its mission to ensure that institutions deliver comparable and consistent radiation doses to their patients, conducts on-site dosimetry review visits to institutions. As a part of the on-site audit, the RPC measures the small field size output factors as might be used in IMRT treatments, and compares the resulting field size dependent output factors to values calculated by the institution's treatment planning system (TPS). The RPC has gathered multiple small field size output factor datasets for X-ray energies ranging from 6 to 18 MV from Varian, Siemens and Elekta linear accelerators. These datasets were measured at 10 cm depth and ranged from 10 × 10 cm(2) to 2 × 2 cm(2). The field sizes were defined by the MLC and for the Varian machines the secondary jaws were maintained at a 10 × 10 cm(2). The RPC measurements were made with a micro-ion chamber whose volume was small enough to gather a full ionization reading even for the 2 × 2 cm(2) field size. The RPC-measured output factors are tabulated and are reproducible with standard deviations (SD) ranging from 0.1% to 1.5%, while the institutions' calculated values had a much larger SD range, ranging up to 7.9% [corrected].The absolute average percent differences were greater for the 2 × 2 cm(2) than for the other field sizes. The RPC's measured small field output factors provide institutions with a standard dataset against which to compare their TPS calculated values. Any discrepancies noted between the standard dataset and calculated values should be investigated with careful measurements and with attention to the specific beam model.

A novel polymerase ß inhibitor from phage displayed peptide library augments the anti-tumour effects of temozolomide on colorectal cancer.

The therapeutic efficacy of TMZ, a common used drug for chemotherapy, is limited by the resistance from colorectal cancer cells. Base excision repair (BER) pathway has been identified as one of the reasons for drug resistance. By blocking Polß-dependent BER (Base Excision Repair) pathway, the efficacy of TMZ treatment can be improved greatly. Several Polß inhibitors that have been identified could not become approved drugs due to lack of potency or specificity. To find therapeutic candidates with exquisite specificity and high affinity to Polß, phage display technology was used in the current research. We screened out a candidate Polß inhibitor, 10 D, that can inhibit the activity of Polßand SP-BER (Short-Patch Base excision Repair) pathway. Co-treatment with 10 D enhanced the sensitivity of colorectal cancer (CRC) cells to TMZ both in vitro and in vivo. Our data suggested that the novel Polß inhibitor we identified can improve TMZ efficacy and optimize CRC chemotherapy.

Comparative study of topical 5-aminolevulinic acid photodynamic therapy (5-ALA-PDT) and surgery for the treatment of high-grade vaginal intraepithelial neoplasia.

OBJECTIVE: To compare the clinical efficacy and safety of 5-aminolevulinic acid photodynamic therapy (ALA-PDT) and local surgical excision in the treatment of high-grade vaginal intraepithelial neoplasia (VAIN). METHODS: This was a retrospective study of a single center. Patients with biopsy-proven high-grade VAIN were counseled on the option of excision or ALA-PDT. After treatment patients were followed up at 3, 6 and 12 months and every 6 months afterwards. Clinical data of two groups were analyzed. RESULTS: Sixty patients were treated with ALA-PDT and 40 patients underwent excision surgery. The clinical characteristics of patients in the two groups were similar. The mean ages of patients treated with ALA-PDT and local surgical excision were 41.93+13.08 (18-70) and 43.25+11.64 (22-61) years old, respectively. The treatment times of PDT for patients in the ALA-PDT group were 5.48±1.41 (2-8) times. At 3-6 months after treatment, the overall complete remission (CR) rate was 93.33% (56/60) in PDT group and 82.5% (33/40) in local surgical excision group. In PDT group, a 12- and 24-month follow-up, the total HPV clearance rate was 68.97% and 77.55%, respectively. At 12-month and 2-year follow-up after local surgical excision, the total HPV clearance rate was 60% and 64.52%, respectively. Further analysis showed that the clinical efficacy was comparable between the two treatments. Patients over 45 years old exhibited a lower HPV clearance rate (P<0.05) than those under 45 years old. It was also observed that persistent HPV infection is a major risk factor for persistent disease. Further comparative analyses showed no significant difference in the CR rate and treatment times between VAINII and VAINIII in ALA-PDT group. Moreover, 4 patients (11.36%) developed recurrent lesions during follow-up after ALA-PDT, whereas 3 patients (8.33%) in the local excision group developed recurrent lesions. In addition, there was no significant difference in the recurrent rate between two groups. CONCLUSIONS: ALA-PDT shows similar efficacy and less side effects compared to local surgical excision.

Exosomes Derived From Hypoxia-Conditioned Stem Cells of Human Deciduous Exfoliated Teeth Enhance Angiogenesis via the Transfer of let-7f-5p and miR-210-3p.

Physiological root resorption of deciduous teeth is a normal phenomenon. How the angiogenesis process is regulated to provide adequate levels of oxygen and nutrients in hypoxic conditions when the dental pulp tissue is reduced at the stage of root resorption is not fully understood. In this study, we designed hypoxic preconditioning (2%) to mimic the physiological conditions. We isolated exosomes from hypoxic-preconditioned SHED (Hypo-exos) cells and from normally cultured SHED cells (Norm-exos). We found that treatment with Hypo-exos significantly enhanced the growth, migration and tube formation of endothelial cells in vitro compared with Norm-exos. We also performed matrigel plug assays in vivo and higher expression of VEGF and higher number of lumenal structures that stained positive for CD31 were found in the Hypo-exos treated group. To understand the potential molecular mechanism responsible for the positive effects of Hypo-exos, we performed exosomal miRNA sequencing and validated that Hypo-exos transferred both let-7f-5p and miR-210-3p to promote the tube formation of endothelial cells. Further study revealed that those two miRNAs regulate angiogenesis via the let-7f-5p/AGO1/VEGF and/or miR-210-3p/ephrinA3 signal pathways. Finally, we found that the increased release of exosomes regulated by hypoxia treatment may be related to Rab27a. Taking these data together, the present study demonstrates that exosomes derived from hypoxic-preconditioned SHED cells promote angiogenesis by transferring let-7f-5p and miR-210-3p, which suggests that they can potentially be developed as a novel therapeutic approach for pro-angiogenic therapy in tissue regeneration engineering.

Therapeutic effects of topical photodynamic therapy with 5-aminolevulinic acid on cervical high-grade squamous intraepithelial lesions.

BACKGROUND: Topical photodynamic therapy with 5-aminolevulinic acid (ALA-PDT) is a non-invasive option for treatment of Cervical Intraepithelial Neoplasia (CIN). We investigated the clinical efficacy and safety of ALA-PDT in treatment of patients with cervical high-grade squamous intraepithelial lesion (HSIL) and evaluated the factors that effect the efficacy of PDT. METHODS: A total of 99 patients (average age: 30 years (18-45 years old)) with cervical HSIL and high-risk Human Papilloma Virus (hr-HPV) infections and who had received ALA-PDT were enrolled in this retrospective study. Before ALA-PDT, cervical exfoliated cells from all patients were obtained for HPV genotyping, liquid-based cytology test (LBC) and PAX1 methylation (PAX1m) test. Then, 6 months after ALA-PDT, HPV genotyping, LBC and colposcopy-directed biopsy were conducted. During subsequent follow-up, HPV genotyping and LBC were performed, and colposcopy-directed biopsy was conducted if hr-HPV was positive and/or LBC result was abnormal. RESULTS: At 6 months after treatment, total HPV clearance and lesions complete remission (CR) rates were 64.6% (64/99) and 88.9% (88/99), while at 1-year of follow-up, they were 81.3% (65/80) and 92.5% (74/80), respectively. At 1-year follow-up, HPV clearance rate was higher, relative to 6 months after ALA-PDT (p <0.05). At 6 months after treatment, HPV clearance and CR rates in the PAX1 hypermethylation (PAX1hm) group were 36.8% and 73.7%, which were lower compared to the rates for PAX1 hypomethylation (PAX1lm) group (71.3%, 92.5%) (p <0.05). Moreover, of the 11 patients with cervical canal lesions, 3 (3/7, 42.9%) patients had cervical canal HSIL persistence and the other 4 cervical canal HSIL and all of the 4 cervical canal LSIL had complete remission. CONCLUSIONS: ALA-PDT is an effective, safe treatment approach for patients with cervical HSIL and hr-HPV infections. Patients with PAX1lm showed better therapeutic outcomes than patients with PAX1hm or cervical canal HSIL.

The effect of local photodynamic therapy with 5-aminolevulinic acid in the treatment of vaginal intraepithelial lesions with high-risk HPV infection.

BACKGROUND: Recent data has shown increased rate of vaginal intraepithelial neoplasia (VAIN) in younger individuals. Traditional VAIN treatments exert adverse effects on the anatomy and functions of the vagina. Topical 5-aminolevulinic acid photodynamic therapy (ALA-PDT) is a novel, non-invasive targeted therapy for intraepithelial lesions. Here, we aimed to investigate the efficacy and safety of ALA-PDT in VAIN with high-risk HPV (HR-HPV) infection. METHODS: This study enrolled a total of 82 HR-HPV-positive women diagnosed with VAIN, who were receiving ALA-PDT. The effect of ALA-PDT was evaluated using HPV genotyping and Thinprep cytologic test (TCT). Besides, colposcopy directed biopsies were performed in all patients at 3-month follow-up, and in patients with positive HR-HPV and/or abnormal TCT results during the follow-up period. RESULTS: Our data showed that the HPV clearance rate was 53.7% (44/82) at 6-month follow-up and 67.1% (55/82) at 12-month follow-up. Patients with HPV16/18 combined with other HR-HPV (HPV16/18 & other HR-HPV) infection, those with a history of hysterectomy and/or older age were had significantly lower HPV clearance rate. Besides, the lesion complete remission (CR) rate was 90.2% (74/82). These results show that HPV16/18 & other HR-HPV infection was associated with significantly lower CR rates. Throughout the follow-up, 2 cases had persistent lesions (2.4%), 6 cases had partial remission (7.3%), 2 cases recurred (2.7%) but none of the patients had disease progression. CONCLUSIONS: Taken together, our data showed that ALA-PDT is an effective, safe, and alternative treatment for VAIN patients with HR-HPV infection. However, the ALA-PDT efficacy was relatively poor in patients with older age, HPV16/18 & other HR-HPV infection and/or those with a history of hysterectomy.