BACKGROUND:The cryopreservation technology enables tissues/cells to be stored for a long time in a low-temperature environment while maintaining the integrity of their activity and function,which is of great significance for the construction of cell therapy,tissue engineering and biological sample banks.Cryoprotective agents often contain dimethyl sulfoxide and serum.To avoid the toxic side effects of dimethyl sulfoxide,the complexity of serum components and immune responses,although some finished cryoprotective agents have been marketed,they are faced with many difficulties such as high cost and limited application.Therefore,there is an urgent need to develop a cryoprotective agent with clear components and the ability to solve the above problems.OBJECTIVE:To evaluate the effects of a novel cryoprotectant on cryopreservation efficiency of different tissue and cell sources.METHODS:By applying the novel cryoprotectant as an experimental group with the commercially available and widely used cryoprotectant(control group)to umbilical cord Wharton's jelly tissue,umbilical cord mesenchymal stem cells,umbilical cord blood/peripheral blood mononuclear cells,NK and CIK cells,comparative analyses were conducted in terms of cell morphology,number,viability,surface markers,differentiation potential,and cell-killing toxicity assay before cryopreservation and after resuscitation thawing.We confirmed the cryopreservation effect of the new cryoprotectant and its potential application value.RESULTS AND CONCLUSION:(1)The novel cryoprotectant facilitated the normal growth of cryopreserved Wharton's jelly tissue upon recovery,exhibiting mesenchymal stem cell morphology.No significant differences were observed between the experimental and control groups in terms of cell recovery rate,surface markers,and differentiation potential.(2)There was no significant difference in the number and viability of cells between the experimental group and the control group after cryopreservation of cord blood/peripheral blood mononuclear cells,and the cryo-resuscitated cell numbers and viability of derived NK cells/CIK cells did not show significant difference between the experimental and control groups.(3)For NK cells derived and differentiated from cord blood/peripheral blood mononuclear cells,there was no significant difference in the proportion of CD56+CD16+cell subpopulations between the experimental group and the control group.For CIK cells derived and differentiated from cord blood/peripheral blood mononuclear cells,there was no significant difference in the proportions of CD3+CD8+and CD3+CD56+cell subpopulations between the experimental group and the control group.(4)In terms of cytotoxicity testing,when the effective-target ratio of immune cells and melanoma cell line Mel624 was 20:1,whether it was NK cells/CIK cells derived from cord blood or peripheral blood mononuclear cells,there was no significant difference in the tumoricidal activity of cells between the experimental group and the control group.These findings suggest that the novel cryoprotectant can replace existing commercially available and widely used cryoprotectants,and is applicable to Wharton's jelly tissue,umbilical cord mesenchymal stem cells,umbilical cord blood/peripheral blood mononuclear cells,as well as NK and CIK cells,providing a solid technical foundation for the scaling,standardization,and commercialization of universal cryoprotectants.

Objective:To investigate the association between serum 25-hydroxy vitamin D [25-(OH)D] levels at 1 and 4 weeks after birth and bronchopulmonary dysplasia (BPD) in preterm infants.Methods:This case-control study included 125 preterm infants (gestational age<32 weeks) admitted to the Department of Neonatology, Hangzhou Women's Hospital between January 2020 and December 2022. All infants received vitamin D supplementation (900 IU/d) starting at 1 week after birth. According to the clinical diagnosis at 28 d after birth, these participants were divided into BPD ( n=57) and non-BPD ( n=68) groups. Two independent sample t-test, Wilcoxon rank-sum test, and Chi-square test were used to compare the general conditions, maternal perinatal conditions, postnatal treatment, complications, and 25-(OH)D levels between the two groups. Pearson bivariate correlation and partial correlation analyses were performed to assess the relationship between 25-(OH)D levels and BPD. Results:The BPD group had lower gestational age [(28.7±1.8) vs. (30.2±0.9) weeks, t=－6.13], birth weight [(1 166.1±238.1) vs. (1 473.8±224.4) g, t=－7.42], and Apgar scores at 1 min [(7.6±2.1) vs. (9.1±1.3) scores, t=－4.58) and 5 min [(8.9±1.2) vs. (9.5±1.1) scores, t=－2.77) compared to the non-BPD group (all P<0.05). The proportion of infants received surfactant therapy [73.7% (42/57) vs. 50.0% (34/68), χ2=7.30], the incidence of maternal preeclampsia [29.8% (17/57) vs. 14.7% (10/68), χ2=4.19], and the incidence of neonatal asphyxia [31.6% (18/57) vs. 16.2% (11/68), χ2=4.13], pneumonia [14.0% (8/57) vs. 1.5% (1/68), χ2=5.57], sepsis [64.9% (37/57) vs. 19.1% (13/68), χ2=27.10], and patent ductus arteriosus [78.9% (45/57) vs. 48.5% (33/68), χ2=12.23] were higher in the BPD group than in the non-BPD group (all P<0.05). The infants with BPD required longer durations of caffeine therapy [(37.2±15.1) vs. (16.8±11.5) d, t=8.58], mechanical ventilation [0.0 (0.0-3.0) vs. 0.0 (0.0-0.0) d, Z=3.52], non-invasive ventilation [23.0 (11.0-31.0) vs. 6.0 (4.0-9.0) d, Z=6.22], total oxygen therapy [46.0 (40.0-58.0) vs. 13.0 (6.0-19.0) d, Z=6.57], and hospitalization [(60.7±15.0) vs. (37.6±7.8) d, t=10.52] than those without BPD (all P<0.05). Serum 25-(OH)D levels were significantly lower in the BPD group than in the non-BPD group at both 1 week [(32.75±7.81) vs. (43.07±9.36) nmol/L, t=－4.60, P<0.001] and 4 weeks [(49.03±11.12) vs. (60.02±14.39) nmol/L, t=－3.90, P<0.001] after birth. Vitamin D deficiency at 1 week after birth was more prevalent in the BPD group than in the non-BPD group [40.0% (10/25) vs. 12.5% (5/40), χ2=6.55, P=0.010]. Serum 25-(OH)D levels at 1 and 4 weeks after birth were negatively correlated with BPD incidence (bivariate analysis: r=－0.50 and－0.40; partial correlation analysis corrected with birth gestational age, birth weight and other general information, the proportion of preeclampsia/cesarean section and other perinatal information of pregnant women, sepsis/mechanical ventilation time and other postnatal diseases and treatment conditions: r=－0.37 and－0.27; both P<0.05). Conclusion:Low serum 25-(OH)D levels at 1 and 4 weeks after birth may be associated with BPD in preterm infants.

BACKGROUND:The cryopreservation technology enables tissues/cells to be stored for a long time in a low-temperature environment while maintaining the integrity of their activity and function,which is of great significance for the construction of cell therapy,tissue engineering and biological sample banks.Cryoprotective agents often contain dimethyl sulfoxide and serum.To avoid the toxic side effects of dimethyl sulfoxide,the complexity of serum components and immune responses,although some finished cryoprotective agents have been marketed,they are faced with many difficulties such as high cost and limited application.Therefore,there is an urgent need to develop a cryoprotective agent with clear components and the ability to solve the above problems.OBJECTIVE:To evaluate the effects of a novel cryoprotectant on cryopreservation efficiency of different tissue and cell sources.METHODS:By applying the novel cryoprotectant as an experimental group with the commercially available and widely used cryoprotectant(control group)to umbilical cord Wharton's jelly tissue,umbilical cord mesenchymal stem cells,umbilical cord blood/peripheral blood mononuclear cells,NK and CIK cells,comparative analyses were conducted in terms of cell morphology,number,viability,surface markers,differentiation potential,and cell-killing toxicity assay before cryopreservation and after resuscitation thawing.We confirmed the cryopreservation effect of the new cryoprotectant and its potential application value.RESULTS AND CONCLUSION:(1)The novel cryoprotectant facilitated the normal growth of cryopreserved Wharton's jelly tissue upon recovery,exhibiting mesenchymal stem cell morphology.No significant differences were observed between the experimental and control groups in terms of cell recovery rate,surface markers,and differentiation potential.(2)There was no significant difference in the number and viability of cells between the experimental group and the control group after cryopreservation of cord blood/peripheral blood mononuclear cells,and the cryo-resuscitated cell numbers and viability of derived NK cells/CIK cells did not show significant difference between the experimental and control groups.(3)For NK cells derived and differentiated from cord blood/peripheral blood mononuclear cells,there was no significant difference in the proportion of CD56+CD16+cell subpopulations between the experimental group and the control group.For CIK cells derived and differentiated from cord blood/peripheral blood mononuclear cells,there was no significant difference in the proportions of CD3+CD8+and CD3+CD56+cell subpopulations between the experimental group and the control group.(4)In terms of cytotoxicity testing,when the effective-target ratio of immune cells and melanoma cell line Mel624 was 20:1,whether it was NK cells/CIK cells derived from cord blood or peripheral blood mononuclear cells,there was no significant difference in the tumoricidal activity of cells between the experimental group and the control group.These findings suggest that the novel cryoprotectant can replace existing commercially available and widely used cryoprotectants,and is applicable to Wharton's jelly tissue,umbilical cord mesenchymal stem cells,umbilical cord blood/peripheral blood mononuclear cells,as well as NK and CIK cells,providing a solid technical foundation for the scaling,standardization,and commercialization of universal cryoprotectants.

Objective:To investigate the association between serum 25-hydroxy vitamin D [25-(OH)D] levels at 1 and 4 weeks after birth and bronchopulmonary dysplasia (BPD) in preterm infants.Methods:This case-control study included 125 preterm infants (gestational age<32 weeks) admitted to the Department of Neonatology, Hangzhou Women's Hospital between January 2020 and December 2022. All infants received vitamin D supplementation (900 IU/d) starting at 1 week after birth. According to the clinical diagnosis at 28 d after birth, these participants were divided into BPD ( n=57) and non-BPD ( n=68) groups. Two independent sample t-test, Wilcoxon rank-sum test, and Chi-square test were used to compare the general conditions, maternal perinatal conditions, postnatal treatment, complications, and 25-(OH)D levels between the two groups. Pearson bivariate correlation and partial correlation analyses were performed to assess the relationship between 25-(OH)D levels and BPD. Results:The BPD group had lower gestational age [(28.7±1.8) vs. (30.2±0.9) weeks, t=－6.13], birth weight [(1 166.1±238.1) vs. (1 473.8±224.4) g, t=－7.42], and Apgar scores at 1 min [(7.6±2.1) vs. (9.1±1.3) scores, t=－4.58) and 5 min [(8.9±1.2) vs. (9.5±1.1) scores, t=－2.77) compared to the non-BPD group (all P<0.05). The proportion of infants received surfactant therapy [73.7% (42/57) vs. 50.0% (34/68), χ2=7.30], the incidence of maternal preeclampsia [29.8% (17/57) vs. 14.7% (10/68), χ2=4.19], and the incidence of neonatal asphyxia [31.6% (18/57) vs. 16.2% (11/68), χ2=4.13], pneumonia [14.0% (8/57) vs. 1.5% (1/68), χ2=5.57], sepsis [64.9% (37/57) vs. 19.1% (13/68), χ2=27.10], and patent ductus arteriosus [78.9% (45/57) vs. 48.5% (33/68), χ2=12.23] were higher in the BPD group than in the non-BPD group (all P<0.05). The infants with BPD required longer durations of caffeine therapy [(37.2±15.1) vs. (16.8±11.5) d, t=8.58], mechanical ventilation [0.0 (0.0-3.0) vs. 0.0 (0.0-0.0) d, Z=3.52], non-invasive ventilation [23.0 (11.0-31.0) vs. 6.0 (4.0-9.0) d, Z=6.22], total oxygen therapy [46.0 (40.0-58.0) vs. 13.0 (6.0-19.0) d, Z=6.57], and hospitalization [(60.7±15.0) vs. (37.6±7.8) d, t=10.52] than those without BPD (all P<0.05). Serum 25-(OH)D levels were significantly lower in the BPD group than in the non-BPD group at both 1 week [(32.75±7.81) vs. (43.07±9.36) nmol/L, t=－4.60, P<0.001] and 4 weeks [(49.03±11.12) vs. (60.02±14.39) nmol/L, t=－3.90, P<0.001] after birth. Vitamin D deficiency at 1 week after birth was more prevalent in the BPD group than in the non-BPD group [40.0% (10/25) vs. 12.5% (5/40), χ2=6.55, P=0.010]. Serum 25-(OH)D levels at 1 and 4 weeks after birth were negatively correlated with BPD incidence (bivariate analysis: r=－0.50 and－0.40; partial correlation analysis corrected with birth gestational age, birth weight and other general information, the proportion of preeclampsia/cesarean section and other perinatal information of pregnant women, sepsis/mechanical ventilation time and other postnatal diseases and treatment conditions: r=－0.37 and－0.27; both P<0.05). Conclusion:Low serum 25-(OH)D levels at 1 and 4 weeks after birth may be associated with BPD in preterm infants.

Tumor suppressor gene O⁶-methylguanine-DNA methyltransferase (MGMT) promoter methylation has been reported in melanoma. However, the clinical and prognostic significance of MGMT promoter methylation in patients with melanoma remained to be determined. A systematic search was performed to identify eligible papers published. The overall odds ratios (ORs) or hazard ratios and their 95% confidence intervals were calculated. Final 12 eligible publications involving Caucasian population were performed in this study, including 1,071 metastatic melanoma patients, 154 primary melanoma patients, and 211 normal controls. MGMT promoter methylation was significantly higher in primary or metastatic melanoma than in normal controls (p < 0.05). No difference of MGMT promoter methylation was found in primary and metastatic melanoma (p=0.432). When metastatic melanoma was compared to normal controls, subgroup analysis showed the correlation between MGMT promoter methylation and different sample materials (tissue: OR=7.01, p < 0.001 and blood: OR=12.04, p=0.005). MGMT promoter methylation was not associated with response to drug therapy and the prognosis in overall survival and progression-free survival for multivariate analysis. Our results show that MGMT promoter methylation may be correlated with the increased risk of primary or metastatic melanoma. Based on blood samples, MGMT promoter methylation may become a noninvasive biomarker for the detection of metastatic melanoma. Further additional clinical studies are necessary.
Disease-Free Survival ; Drug Therapy ; Genes, Tumor Suppressor ; Humans ; Melanoma* ; Methylation* ; Multivariate Analysis ; Odds Ratio ; Prognosis

OBJECTIVETo observe the histological features of tumor-bearing tissues formed by human fibroblasts after reprograming by spermatogonial stem cell self-renewal key regulating gene Piwil2 (Piwil2-iCSC).

METHODSPiwil2-iCSC tumor spheroids-like colonies were selected for tumor formation assay in four nude mice. Pathological features of Piwil2-iCSC tumors were observed by histology. Stem cell markers and common triploblastic markers were detected by reverse transcriptase-polymerase chain reaction (RT-PCR) assay and immunohistochemistry. Germ cell tumor markers were detected by immunohistochemical examination.

RESULTSTwo weeks after inoculation, subcutaneous tumors were formed in all the four nude mice with a tumor formation rate of 100%. In the Piwil2-iCSC tumor tissues, Piwil2-GFP(+) cells showed high-density nuclear expression and were widely observed in DAPI-stained sections. Numerous mitotic figure of the neoplastic cells were seen (>10 cells/field of vision under high magnification) in HE-stained sections. Enlarged abnormal cell nuclei were observed. RT-PCR assay showed that Piwil2-iCSC tumors still expressed Piwil2 and some self-renewal and pluripotent markers of stem cells and some markers of triploblastic differentiation. Immunohistochemical staining showed that the tumors expressed stem cell markers, triploblastic markers and germ cell tumor markers AFP and HCG.

CONCLUSIONSPiwil2-iCSC tumors are probably undifferentiated embryonic small cell carcinoma, most likely to be immature teratoma, mixed with yolk sac tumor and choriocarcinoma components. It can be used as a useful model for the research of origin or genesis mechanism of cancer stem cells and the treatment of relevant tumors.

Adult Stem Cells ; Animals ; Argonaute Proteins ; genetics ; Cellular Reprogramming Techniques ; Choriocarcinoma ; pathology ; Endodermal Sinus Tumor ; pathology ; Fibroblasts ; metabolism ; pathology ; Humans ; Immunohistochemistry ; Mice ; Mice, Nude ; Neoplasms, Germ Cell and Embryonal ; chemistry ; genetics ; pathology ; Neoplastic Stem Cells ; chemistry ; pathology ; Real-Time Polymerase Chain Reaction ; Spheroids, Cellular ; Teratoma ; pathology ; Time Factors

Objective To explore the clinicopathologic features of pityriasis lichenoides et varioliformis acuta (PLEVA).Methods A retrospective analysis was performed.Clinical and histological data were collected from 60 patients with PLEVA.The clinicopathologic features of PLEVA were analyzed.Results Among the 60 patients with PLEVA,32 (53.3％) were aged 2-18 years,and 28 (46.7％) aged 19-49 years.Skin lesions were distributed in a diffuse pattern in 50 (83.3％) patients,in a central pattern in 2 (3.3％) patients,and in a peripheral pattern in 8 (13.4％) patients.Nineteen (31.6％) patients had a history of upper respiratory infection.Histopathological examination revealed liquefactive degeneration of basal cells and perivasculitis in the dermis in all the 60 cases,neutrophil abscess formation in the stratum corneum in 26 (43.3％) cases,keratinocyte necrosis in the epidermis in 41 (68.3％) cases,generalized liquefactive degeneration in 30 (50.0％) cases,migration of lymphocytes into the epidermis in 43 (71.6％) cases,Pautrier's microabscess formation in 2 cases,varying degrees of extravasation of erythrocytes into the epidermis in 46 (76.7％) cases,fibrinoid necrosis of blood vessel walls in the dermis in 3 cases.PLEVA progressed into granuloma fungoides in 1 patient.Twenty patients underwent immunohistochemical examination,and 3 of them showed monoclonal hyperplasia of T cells.Conclusions PLEVA has characteristic clinical manifestations,and the combination of pathological and clinical examination is the gold standard for its diagnosis.

Objective To explore the effect of health belief model on compliance behavior of patients with PICC(peripherally inserted central catheter).Methods According to admission order,75 cases of PICC were divided into control group(n=36)and observation group(n=39).Patients in the control group received conventional health education and regular visits after discharge and besides the regular methods,patients in the observation group were intervened by health belief model for the compliance behavior.All patients of two groups were investigated with self-designed scale for compliance behavior.Result The compliance behavior of the observation group was better than that of control group with significant difference(P<0.05).Conclusion The health belief model of education can improve the compliance behavior of PICC patients after discharge from hospital.

Objective To study the clinical effect of continuous central venous pressure monitoring and select a better method for central venous pressure momtoring. Methods Continuous central venous pressure wag monitored by connecting pressure sensor to central venous catheter in 56 patients with open heart operation.At the same time routine monitoring method was used in the same central venous vein of the saine patient The CVP values from the two methods were compared for 100 times and analyzed the difference.Results The VCP values of the two groups had no statistical difference(P＞0.05).Conclusions Continuous monitoring by pressure sensor had advantages such as continuous data,dynamic,direct-viewing,Veracious and decreased chance of infection.It could reduce the workload of nurses and possessed more clinical value compared with routine monitoring method.