[Experimental studies on the repair and restitution of cartilage by cartilage acellular extracellular matrix and adipose tissue-derived stem cells].

Objective: To investigate the effects of the repair and restitution of ear-shaped cartilage by adipose tissue-derived stem cells(ADSC) and cartilage acellular extracellular matrix. Methods: ADSC were extracted by digesting with collagenase type II from the adipose tissue from 32 patients with adiposity whose fats were drawn, and were cultured and subcultured in vitro. The natural biological scaffolds were prepared by acellular method using porcine ear cartilage, and then the second generation ADSC(5.0×10(7)/ml) were inoculated on the preformed natural bio-scaffold scaffold by culturing in vitro for 3 days to form a cell scaffold complex. 32 New Zealand white rabbits were randomly divided into the experimental groups, the control group A, the control group B and the control group C. All New Zealand white rabbits were modeled by ear cartilage defects. The cell scaffolds composite was implanted into the experimental group of the ear cartilage defects of rabbits, the ADSC were implanted into the control group A, the cartilage acellular extracellular matrix scaffold was implanted into the control group B and the control group C was modeled only by ear cartilage defects. After 16 weeks, the animals were sacrificed and the repair effect was observed by gross appearance and histological examinations including HE, Toluidine blue staining, Safranin O and typeⅡ collagen staining. Its were quantitatively analyzed by positive staining results of type Ⅱ collagen. Ear cartilage tissue elasticity was detected. SPSS 17.0 software was used to analyze the data. Results: The cartilage defects in the experimental group were repaired well by general shape observation and those in the control group was filled in with granulation tissue. There were significant differences between the experimental group and the control group in the wet weight(P<0.05). HE staining showed that cartilage cavities formed in articular cartilage defects, and only the fibrous tissue was filled with the ear cartilage defect in the control groups. In the repair area, Toluidine blue staining, Safranin O and type Ⅱ collagen staining were positive in the experimental group, and negative in the control groups. There was no significant difference between the experimental group and the normal ear cartilage in the ear cartilage elastic constant detection(P>0.05). Conclusions: The mechanics and histology of rabbit ear neonatal cartilage constructed by ADSC combined with cartilage acellular matrix are close to normal ear cartilage. Cartilage acellular matrix material combined with adipose-derived stem cells has good repair and reconstruction ability for ear cartilage defects, which possesses potential clinical application value.

Developmental exposure to lead causes inherent changes on voltage-gated sodium channels in rat hippocampal CA1 neurons.

In this study, the effects of chronic lead (Pb(2+)) exposure, during day 0 of gestation (E0) to postnatal day 15 (P15), on voltage-gated sodium channel currents (I(Na)) were investigated in CA1 field of the hippocampus (CA1) neurons using the conventional whole-cell patch-clamp technique on rat hippocampal slices. We found that developmental lead exposure increased the activation threshold and the voltage at which the maximum I(Na) current was evoked, caused positive shifts of I(Na) steady-state activation curve, and enlarged I(Na) tail-currents; Pb(2+) delayed the activation of I(Na) in a voltage-dependent manner, prolonged the time course of the fast inactivation of sodium channels; Pb(2+) induced a right shift of the steady-state inactivation curve, accelerated the activity-dependent attenuation of I(Na), but made no significant effects on the time course of the recovery of I(Na) from inactivation and the fraction of inactivated channels. In addition, the co-treatment with alpha-tocopherol (VE), an effective antioxidant and free radical scavenger, completely prevented the aforementioned changes on I(Na). The alterations on I(Na) properties induced by developmental lead exposure were partly different from that in previous acute experiments under the conditions closer to physiological situation, and the process was considered related to the participating of lead in lipid peroxidation reaction, which has been reported to change the conformation and biophysical functions of membrane proteins.

Differential gene expression profiling in aggressive bladder transitional cell carcinoma compared to the adjacent microscopically normal urothelium by microdissection-SMART cDNA PCR-SSH.

Identifying novel and known genes that are differentially expressed in aggressive bladder transitional cell carcinoma (BTCC) has important implications in understanding the biology of bladder tumorigenesis and developing new diagnostic and therapeutic agents. In this study we identified the differential gene expression profiles comparing tumor to the adjacent microscopically normal mucosa by manual microdissection on frozen sections. The RNAs extracted from microdissected tissues were amplified by SMART cDNA PCR technology to generate forward subtractive cDNA library by suppressive subtractive hybridization (SSH). We obtained 376 positive clones, one hundred clones of aggressive BTCC subtracted cDNA library were selected at random and inserts were reamplified by PCR. After differential screening by reverse dot blotting, 73 positive clones, that contend inserts putatively upregulated in aggressive BTCC, were further analysed by DNA sequencing, GenBank and EST database searching. Sequencing results showed that 66 clones stand for 23 known genes and 7 clones for three new EST (Genbank number: DN236875, DN236874 and DN236873). In conclusion, microdissection-SMART cDNA PCR-SSH allowed for an efficient way to identify aggressive BTCC-specific differential expressed genes that may potentially be involved in the carcinogenesis and/or progression of aggressive BTCC. These differentially expressed genes may be of potential utility as therapeutic and diagnostic targets for aggressive BTCC.

Decline of serum levels of free testosterone in aging healthy Chinese men.

OBJECTIVE: To investigate the age-related change of serum androgen levels in healthy men and to define a cut-off value of serum testosterone for the diagnosis of androgen deficiency in the aging male. METHOD: 1080 healthy men aged 20 to 70 years old were enrolled in Beijing, Shanghai, Xian and Chongqing. Luteinizing hormone (LH), follicle-stimulating hormone (FSH), total testosterone (T), calculated free testosterone (cFT), sex hormone binding globulin (SHBG), 17beta-oestradiol (E2), the T/LH ratio, and T/SHBG as a free testosterone index (FTI) were all determined. RESULTS: Serum total T did not significantly decline, but the cFT, T/LH and FTI progressively decreased with aging. To determine androgen deficiency, the 10th percentile value of men < 40 years was defined as the lower cut-off value for cFT, T/LH or FTI, which were 0.3 nmol/L, 2.8 nmol/IU, and 0.4 nmol/IU respectively. With the median value of cFT of men aged between 20 and 49 years as the criterion, the level of cFT was lower in 2.82% of men from 40 to 49 years, in 19.53% from 50 to 59 years, in 22.57% from 60 to 69 years, and in 33.19% of men > or = 70 years. Taking the above value of cFT as the cut-off point, the prevalence of androgen deficiency in men 40-49 years was 13.0%, 31.8% in men 50-59 years, 30.1% in men 60 to 69 years, and 46.7% in men > 70 years. CONCLUSIONS: (i). While serum total T values do not decline with aging, the levels of cFT gradually decline with aging; (ii) when using the value of cFT of the 10th percentile of men aged 20 to 39 years as the cut-off point, the prevalence of androgen deficiency was < 15% before the age of 50 years, and about 30% thereafter, approaching 45% after the age of 70 years; and (iii) in this study the values of T/LH paralleled those of cFT closely; therefore, T/LH could serve as a surrogate for cFT.