Expanding the toolset of fluorescent covalent staining of biological samples by labeling carboxylate and phosphate groups.

Recently, fluorescent covalent staining methods have been developed for visualization of anatomical structures in cells and tissues. Coupled with expansion microscopy, these stains revealed various ultrastructural details. However, the covalently stainable chemical groups have been limited to amines, carbohydrates, and thiols. Here, we developed procedures for covalently labeling tissues for carboxylate and phosphate groups, utilizing carbodiimide crosslinker chemistry. In porcine kidney tissues, the carboxylate and phosphate stain provides 1.8-4.8-fold higher signal intensity than those from the three existing stains. In cancer cells, such stain allows 2-8-fold more accurate identification of nucleoli than the amine stain. In expansion microscopy samples, such stain reveals a variety of sub-cellular structures in tissues when combined with the amine stain. Such stain also allows imaging of lipid-based structures in cultured cells. With these advantages, this new covalent staining method further expands the toolset for fluorescent visualization of histology.

A functional reference map of the RNF8 interactome in cancer.

BACKGROUND: RNF8 is an E3 ligase identified as a critical DNA damage-responsive protein. Recently, multiple reports have shown that RNF8 could be used as an important therapeutic target for cancer chemo/radiotherapy. However, the understanding of RNF8 remains limited due to the lack of its interactome reference map and comprehensive analysis of RNF8 in diverse cancers, which underscores the need to map the interactome of RNF8 via high-throughput methods. RESULTS: A two-way identification method based on LC-MS was designed for the identification of the RNF8 interactome with high-specificity. By in silico analysis and in vitro validation, we identified a new reference map of the RNF8 interactome network containing many new targets, such as YBX1, DNMT1, and HDCA1, new biological functions and the gene-disease associations of RNF8. Our results revealed a close relationship between RNF8 and neurodegenerative diseases or tumor-infiltrating immune cells using bulk RNA-seq and scRNA-seq datasets. As a proof of concept of our interactome map, we validated the direct binding between RNF8 and YBX1 and showed that RNF8 catalyzed the ubiquitination of YBX1. These results demonstrated that RNF8 might be a crucial regulator of YBX1. CONCLUSIONS: Our work provides a unique framework for researchers and clinicians who seek to better explore or understand RNF8-regulated biological functions in cancers. This study will hopefully facilitate the rational design and further development of anti-RNF8 therapy in cancers.

3D printing silk-gelatin-propanediol scaffold with enhanced osteogenesis properties through p-Smad1/5/8 activated Runx2 pathway.

The application of 3 D printing technology in tissue engineering has become increasingly important. However, due to the limitations of bio-ink, there are still some remaining problems. For example, the major challenge for ideal bio-ink is to maintain stable 3 D structure and good biocompatibility in the meantime while conventional gels are week and nearly unprintable. So, the development of new bio-ink material with improved rheological and mechanical properties is highly demanded to avoid compromising biocompatibility for tissue engineering. Silk fibroin (SF), a natural degradable polymer, is considered to be a proper material for the preparation of bio-inks. We used SF, gelatin, and polyols as raw materials to fabricate bio-inks and scaffolds. We evaluated the rheological properties and printability of bio-inks with a rotational rheometer and a 3 D printer. The scaffolds were prepared by crosslinking and freeze-drying technologies. The biocompatibility and osteoinductive functions of scaffolds were investigated by evaluating proliferation, osteogenic differentiation and related cell signaling of cultured MC3T3-E1 cells. The results showed that the scaffolds using SF, Gel and propanediol (PG) not only had good rheological properties and storage modulus, but also could better enhance osteogenic specific genes expression mediated by Smad1/5/8 and Runx2 pathways. What is more, morphological characterization showed that α-mem incubation could help scaffold form porous structure on its surface, which could shed a light on a new 3 D bio-printed bone repair scaffold with both naturally emerged and CAD-designed porous structure. Our findings provide a potential biomaterial for the treatment of bone tissue regeneration.

Catalytic reduction of 4-nitrophenol over Ni-Pd nanodimers supported on nitrogen-doped reduced graphene oxide.

Catalytic reduction of toxic 4-nitrophenol to 4-aminophenol over magnetically recoverable nanocatalysts has attracted much attention. Herein, we report a Ni-Pd/NrGO catalyst through the growth of Ni-Pd nanodimers (NDs) on nitrogen-doped reduced graphene oxide (NrGO). The Ni-Pd NDs show a heterogeneous nanostructure with Ni and Pd subparts contacting with each other, remarkably different from the frequently-observed core/shell nanoparticles (NPs) or nanoalloy. The formation of Ni-Pd NDs follows an initial deposition of Pd NPs on the graphene and in-situ catalytic generation of Ni subparts over the newly-generated Pd NPs. The resulting Ni-Pd/NrGO exhibits a superior catalytic activity towards the reduction of 4-nitrophenol at room temperature with a high rate constant (3400s-1g-1) and a low activated energy (29.1kJmol-1) as compared to unsupported Ni-Pd NDs and supported monometallic catalysts. The conversion rate of 4-NP is calculated to be 99.5% and the percent yield (%) of 4-AP is as high as 99.1%. A synergistic catalysis mechanism is rationally proposed, which is ascribed to the electronic modification of Ni-Pd metals due to the strong metal/support interaction (SMSI) effect as well as the electron transfer between Ni and Pd. The hybrid catalyst shows soft ferromagnetic properties and can be magnetically separated and recycled without obvious loss of activity.

Automated feature set selection and its application to MCC identification in digital mammograms for breast cancer detection.

We propose a fully automated algorithm that is able to select a discriminative feature set from a training database via sequential forward selection (SFS), sequential backward selection (SBS), and F-score methods. We applied this scheme to microcalcifications cluster (MCC) detection in digital mammograms for early breast cancer detection. The system was able to select features fully automatically, regardless of the input training mammograms used. We tested the proposed scheme using a database of 111 clinical mammograms containing 1,050 microcalcifications (MCs). The accuracy of the system was examined via a free response receiver operating characteristic (fROC) curve of the test dataset. The system performance for MC identifications was Az = 0.9897, the sensitivity was 92%, and 0.65 false positives (FPs) were generated per image for MCC detection.

Heat shock protein 70 chaperoned alpha-fetoprotein in human hepatocellular carcinoma cell line BEL-7402.

AIM: To investigate the interaction between heat shock protein 70 (HSP70) and ?-fetoprotein (AFP) in human hepatocellular carcinoma (HCC) cell line BEL-7402. METHODS: The expression and localization of HSP70 and AFP in human HCC cell line BEL-7402 were determined by immunocytochemistry and indirect immunofluorescence cytochemical staining. The interaction between HSP70 and AFP in HCC cells was analyzed by immunoprecipitation and Western blot. RESULTS: Immunocytochemical staining detection showed that HCC cell BEL-7402 expressed a high level of HSP70 and AFP synchronously. Both were stained in cell plasma. AFP existed in the immunoprecipitate of anti-HSP70 mAb, while there was HSP70 in the immunoprecipitate of anti-AFP mAb. CONCLUSION: HSP70 chaperones AFP in human HCC cell BEL-7402. The interaction between HSP70 and AFP in human HCC cell can be a new route to study the pathogenesis and immunotherapy of HCC.

Correlation between clinicopathology and expression of heat shock protein 70 and glucose-regulated protein 94 in human colonic adenocarcinoma.

AIM: To investigate the correlation between clinicopathology and expression of heat shock protein 70 (HSP70) and glucose-regulated protein 94 (grp94) in human colonic carcinoma. METHODS: The expression of HSP70 and grp94 was studied in 80 human colonic cancers with or without metastasis as well as in their adjacent mucous membrane by way of immunohistochemistry and pathology photograph analysis. RESULTS: The expression of HSP70 and grp94 was significantly higher in cancer than that in adjacent mucous membrane (92.5%, 85.0% vs 56.3%, 42.5%, P<0.01). HSP70 and grp94 expressed higher in moderately- and poorly-differentiated colonic cancers than that in their adjacent tissues (93.7%, 87.5%; 100%, 90% vs 56.3%, 42.5%; P<0.01). Dukes C and D stages of colonic cancers showed higher positive rates than Dukes A and B stage groups (97.1%, 91.2%; 100%, 90.9%; vs 80%, 70%; 78.6%, 71.4%; P<0.05). There were definite differences in HSP70 and grp94 expression between metastasis groups and non-metastasis groups (100% vs 75%, 100% vs 50%, P<0.05). CONCLUSION: The HSP70 and grp94 expression rates in colonic cancer groups are significantly higher than that in their adjacent mucous membrane. The HSP70 and grp94 expression in poorly-differentiated colonic cancers with metastasis is significantly higher than well-differentiated cancers without metastasis. The overexpression of HSP70 and grp94 can be used as diagnostic or prognostic markers for colonic cancer.

Expression and significance of heat shock protein 70 and glucose-regulated protein 94 in human esophageal carcinoma.

AIM: To investigate the expression and significance of heat shock protein 70 (HSP70) and glucose-regulated protein 94 (grp94) in human esophageal carcinoma and adjacent normal tissues. METHODS: The expression of HSP70 and grp94 in 78 human esophageal cancer and adjacent normal tissues was studied by immunohistochemistry and pathology photograph analysis. RESULTS: Both esophageal cancer and adjacent normal tissues could express HSP70 and grp94. Of the 78 cases of esophageal carcinoma, 95.0%(72/78) showed positive HSP70, mainly stained in nuclei, while grp94 was mainly stained in cell plasma, and the positive rate was 71.8%(56/78). There was a significant difference in the expression of HSP70 and grp94 between esophageal cancer and adjacent normal tissues (P<0.01). Compared with adjacent normal tissues, there was a significant difference between differential types and HSP70 expression (P<0.01). CONCLUSION: HSP70 and grp94 express differently in cell plasma and nuclei. The expression intensity of HSP70 is related to the differentiation of esophageal carcinoma.

[Analysis of fetal distress in pregnancy with hepatitis B virus infection].

OBJECTIVE: To investigate the cause, prognosis and the treatment of fetal distress in pregnant women with hepatitis B virus (HBV) infection. METHODS: eighty one pregnant women and their newborns were selected. The HBV surface antigen (HBsAg), e antigen (HBeAg), core antibody (HBcAb) and deoxyribonucleic acid (HBV DNA) was positive and the hepatic function normal. eighty five pregnant women without HBV infection, normal hepatic function and their newborns were collected as control. The clinical data, serology examination, placenta histological examination and fetal prognosis of the two groups was analysed 76 infants of the investigation group were vaccinated hepatitis B vaccine 10 microgram at 0, 1 and 6 month respectively. Infant's hepatitis B surface antibody (HBsAb) was detected at 24 month. RESULTS: (1) The incidence of the fetal distress in investigation group was 38.3, %, in the control group was 16.5% (P < 0.05). (2) The main reason was chorion angiopathy induced by HBV infection of placenta. (3) In infants with fetal distress, the block rate from mother to fetus was 78.6%, for the infants without distress was 91.7% (P < 0.05). CONCLUSION: HBV infection during pregnancy may cause placenta chorion angiopathy, reduce placenta function, lead to fetal distress and result in immunization failure.