A Simple and Reliable Cooling Approach for the Cryopreservation of Hematopoietic Stem Cells: the Passive Cooling Rate-controlled Technique.

BACKGROUND: Cryopreservation of hematopoietic stem cells (HSCs) has widely been used in stem cell transplantation and cellular therapy treating various human diseases. However, the current conventional cooling approach for the cryopreservation of HSCs has the following potential problems: (1) requirement of a very expensive computer-programmed liquid nitrogen freezer (LNF) for the cooling rate control, (2) a large consumption of liquid nitrogen, (3) periodic breakdown of the LNF due to the mechanical failure of the liquid nitrogen valves (i.e., magnetic-solenoid valves) inside the LNF, and (4) constant monitoring of the LNF operation during the HSCs cooling process. OBJECTIVE: To test and evaluate a simple and reliable approach for the cryopreservation of HSCs using the passive cooling technique. MATERIALS AND METHODS: A passive cooling-rate-controlled device (PCD) was developed and used to cryopreserve HSCs. The PCD is inexpensive, simple, and user-friendly, which needs only the minimum maintenance and no consumption of liquid nitrogen. The PCD was compared to the LNF for the cryopreservation of HSCs in the present study through experiments. The cell viability and functionality were evaluated after cryopreservation. RESULTS: In comparison with the LNF method, the PCD approach enabled high cell viability/survival, recovery rate, and functionality after cryopreservation processes. CONCLUSION: The PCD offers a cost-effective, simple, and reliable approach for the optimal cryopreservation of HSCs.

[A single center large cohort study on perioperative complications of carotid endarterectomy of 547 cases].

Objective: To analyze the risk factors of perioperative complications within 30 days of carotid endarterectomy(CEA) in the treatment of carotid atherosclerosis stenosis(CAS) during 2011-2017, and to discuss the techniques for reducing the perioperative complication rates. Methods: From August 2011 to August 2017, 486 patients with CAS were retrospective included, and 61 of them underwent bilateral CEA, with a total of 547 cases of CEA included. Perioperative complications were collected within 30 days after operation, and the risk factors related to perioperative complications were analyzed by statistical analysis. Results: In total 547 cases, 12 cases had a postoperative stroke, while 1 case died. A total of 7 cases underwent cranial nerve injury, and 5 cases had an incision related complications. In chi-square test analysis, data suggested that there was a significant difference in the incidence of complications in patients with heart disease, preoperative neurological score difference, contralateral carotid serious stenosis or occlusion and intraoperative shunt in CCA/ICA technique application (P<0.05). In the multivariate Logistic regression, it suggested that poor preoperative neurological score and contralateral carotid serious stenosis or occlusion were independent risk factors for perioperative stroke and death. Conclusion: Our results showed that CEA is effective to prevent stroke and treat patients with CAS. Patients with poor preoperative neurological score and contralateral carotid serious stenosis or occlusion may increase the risk of postoperative stroke rates.

Narcissus yellow stripe virus and Narcissus mosaic virus detection in Narcissus via multiplex TaqMan-based reverse transcription-PCR assay.

AIMS: Development of a multiplex TaqMan RT-qPCR assay to simultaneously detect Narcissus yellow stripe virus (NYSV) and Narcissus mosaic virus (NMV), frequently causing mixed narcissus infection. Feasibility verification was confirmed in natural samples. METHODS AND RESULTS: Primers and probes were designed based on the conserved CP gene regions of NYSV or NMV and their suitability for singleplex and multiplex TaqMan RT-qPCR assays as well as for conventional RT-PCR. Conventional RT-PCR, singleplex and multiplex TaqMan RT-qPCR assays proved to be NYSV and NMV specific. P-values and coefficients of variation of TaqMan RT-qPCR assays indicated high reproducibility. Significantly increased sensitivity was achieved compared to conventional RT-PCR. The detection limit of both viruses was 103 copies with superior correlation coefficients and linear standard curve responses between plasmid concentrations and Ct values. NYSV and NMV infection of narcissus leaves, petals and bulbs could successfully be detected via our multiplex RT-qPCR method at 1·25 mg. CONCLUSION: Our multiplex TaqMan RT-qPCR assay provides rapid, specific, sensitive and reliable testing to simultaneously detect NYSV and NMV, supplying useful routine monitoring for different narcissus samples. SIGNIFICANCE AND IMPACT OF THE STUDY: Efficient identification and discrimination of the narcissus viruses provides reliable information for scientists and conventional growers. Furthermore, it enriches the information of NYSV, NMV and other narcissus viruses.

Serum Cyr61 as a potential biomarker for diagnosis of colorectal cancer.

PURPOSE: To determine the sensitivity and specificity of serum Cyr61 as a potential biomarker for the diagnosis of colorectal cancer (CRC) and to assess the association between serum Cyr61 level and CRC clinicopathological status. METHODS: We used an enzyme-linked immunosorbent assay to measure serum Cyr61 in patients with CRC, patients with colorectal adenomas, and healthy controls. We also analyzed the relationship between serum Cyr61 and clinicopathological features of CRC patients. The levels of serum carcinoembryonic antigen (CEA) and carbohydrate antigen 19-9 (CA19-9) were quantified using the Roche Cobas 6000 Analyzer. The sensitivity and specificity of Cyr61, CEA, CA19-9 and CEA + CA19-9 were evaluated by receiver operating characteristic (ROC) analysis. RESULTS: The serum level of Cyr61 was significantly increased in CRC patients compared with colorectal adenoma patients and healthy controls (p < 0.001). Furthermore, the area under the ROC curve for Cyr61 was 0.935 (95 % confidence interval 0.902-0.968), higher than that for CEA + CA19-9 (0.827, 95 % confidence interval: 0.783-0.871). Use of a Cyr61 cutoff value of 92.0 pg/mL allowed distinguishing CRC patients and healthy controls with a sensitivity of 83 % and a specificity of 97 %. Among CRC patients, an elevated level of serum Cyr61 was significantly associated with more advanced TNM stage (p < 0.0042), lymph node metastasis (p < 0.0088), and vascular invasion (p = 0.0027). CONCLUSION: Cyr61 has potential as a serum biomarker for the diagnosis of CRC and for assessment of the clinicopathological status of CRC.

Protein synthesis in the livers of aging mice studied by electron microscopic radioautography.

The application of 3H-leucine results in labeling of the liver cells of mice in which protein is synthesized at various ages of the animals. Quantitative changes of protein synthesis in the hepatocytes of aging mice were studied by electron microscopic radioautography. The silver grains in the hepatocytes were mainly located over the rough surfaced endoplasmic reticulum, mitochondria, Golgi apparatus, cytoplasmic matrix, and a few over the nuclei. The number of silver grains in the cytoplasm and nuclei of the hepatocytes gradually increased after birth, reached the maximum at 1 month after birth, thereafter it continued to decrease with aging until the 24th month. The number of silver grains in the hepatocyte cytoplasm was more than that in nuclei at various ages. The number of silver grains in the rough surfaced endoplasmic reticulum and mitochondria gradually increased from embryo to 1 month after birth, thereafter it continued to decrease with aging until the 24th month. The number of silver grains in the Golgi apparatus showed almost no change from fetal stage to 6 months after birth, thereafter it continued to decrease with aging until the 24th month. The number of silver grains in the cytoplasmic matrix gradually increased from fetal stage to 2 months after birth, then decreased with aging until the 24th month. These changes reflect the quantity of protein synthesized in each cell organelle at various ages of animals.