0.05).IHC loss was found under hypoxia for 2 h to 48 h in a time dependent manner,the cell density in unit area had remarkable difference compared with control(P

Objective To investigate the correlation of cochlear injury and expressions of Bcl-2 and Bax in the cochlea of diabetic rats.Methods Thirty five Wistar rats were randomly divided into diabetic group(n=25) and control group(n=10).Diabetic deafness was induced in rats by intraperitoneal injection of streptozotocin,while the rats in normal group were injected with normal saline.Immunohistochemical staining was used to detect the protein levels of Bcl-2 and Bax in the cochlea after 3 months.Results No significantly morphological change in cochlea of diabetic rats was found.The Bcl-2 levels in stria vascularis in diabetic group and control group were 135.32?21.69 and 45.32?9.08,respectively,there was significant difference(P

BACKGROUND: Studies are very few regarding the specific reaction of bone marrow mesenchymal stem cells (BMSCs) to activated microglia. Moreover, it remains unclear how MSCs maintain dopaminergic neuronal survival under specific microenvironment.OBJECTIVE: To explore the effect of BMSCs stimulated by activated microglia on dopaminergic neuron survival.METHODS: BMSCs were isolated from Wistar rats by attachment method, and in vitro cultured; microglia was activated, and dopaminergic neurons were cultured by enzyme digestion method. The experiment included 5 groups: BMSCs, microglia, lipopolysaccharide (LPS)+microglia; BMSCs+LPS+microglia groups, in which the dopaminergic neurons were cultured with corresponding culture medium; the dopaminergic neurons alone group was cultured with 10% fetal bovine serum+ DMEM/F12. The effect of different microenvironment on dopaminergic neuron survival and gliocyte-derived neurotrophic factor released from BMSCs were detected by immunofluorescence technique.RESULTS AND CONCLUSION: The release of gliocyte-derived neurotrophic factor in groups involving BMSCs was greater than corresponding control group. Tyrosine hydroxylase immunofluorescence showed that neuronal survival of dopaminergic neurons alone group was 15%, microglia group was 10%, LPS+microglia was 5%, but BMSCs+LPS+microglia group was 28%, significantly greater than the other groups (P < 0.05). In addition, survival of in vitro cultured dopaminergic neurons was decreased with increasing culture duration, but the survival of dopaminergic neurons in group involving BMSCs was significantly greater than corresponding control group. This indicates that microglia activation stimulated BMSCs to upregulate gliocyte-derived neurotrophic factor to prevent dopaminergic neurons from toxic injury, and inhibit delayed death of dopaminergic neurons.

Objective To investigate the protective effects of magnesium sulfate on glutamate-induced neurotoxicity in cultured cochlear.Methods The cortis of newborn rats were cultivated in vitro. The cultured cochlear neurons were divided into glutamate group treated with 0.1 mmol?L-1 glutamate,magnesium sulfate group treated with different concentrations of magnesium sulfate(0.5,1.0,2.0 and 4.0 mmol?L-1) and combined group treated with glutamate and magnesium sulfate for 24 h. Immunofluorescence staining of Neurofilament 200kDa antibody was used to label spiral ganglion neuron(SGN) and the terminate of dendrites. The morphological changes of SGN were examined by fluorescent microscope and the amount of SGN was determined. In addition,the SGN was loaded with 10 ?mol?L-1 Fluo-3 /AM,and the intracellular Ca2+was measured by laser confocal scanning microscope (LCSM). Results Many SGN were smaller and swelling and vacuoles at the terminate of dendrites following glutamate treatment for 24 h. In combined group treated with glutamate and 0.5,1.0 and 2.0 mmol?L-1 of magnesium sulfate,the amounts of SGN increased compared with glutamate group(P

Linoleic acid (C18∶2n-6) and ?-linolenic acid (C18∶3n-3) are found widely in fungi, plants and some lower animals. However, they can not be synthesized in mammals due to lack of △12 and ?-3 fatty acid desaturases. To enable endogenous production of essential fatty acids in mammalian cells, here the stable expression of a Caenorhabditis elegans gene FAT-2 encoding △12 fatty acid desaturase in CHO cells was reported. First, the FAT-2 coding sequence was cloned by RT-PCR. To facilitate high level synthesis of heterogeneous protein, the codon usage of the fatty acid desaturase genes was optimized according to the codon preference of mouse by site-directed mutagenesis, 2 synonymous mutations were introduced into FAT-2 gene by overlapping PCR. The codon-modified gene was finally fused to pBudCE4.1 vector (Invitrogen) under the control of CMV promoter. The expression vector pBudCE-FAT2 was linearized with NheⅠ, and then transfected CHO cells, the cells were under Zeocin selection for nine days and then propagated, then the transfected cells were harvested. The genome and total RNA were isolated for PCR and Norhern blot ananlysis. The results revealed that FAT-2 gene has been integrated into the genome of CHO cells and expressed properly. Fatty acids of total cellular lipids were analyzed by gas chromatography. The results indicate that the expression and function of △-12 fatty acid desaturase resulted in accumulation of linoleic acid. The levels of linoleic acid in transgenic cells were 2.4-fold higher than those in wild-type cells. The moderate linoleic acid in CHO cells was derived from cell culture media uptaken by cell membrane. The results demonstrate that a heterogenous desaturase gene can function well in mammalian cells and prove that transgenic approach is an efficient strategy for changing fatty acid composition of mammals.

Vaccination has been proved to be the most effective strategy to prevent the Corona Virus Disease 2019 (COVID-19). The mRNA vaccine based on nano drug delivery system (NDDS) - lipid nanoparticles (LNP) has been widely used because of its high effectiveness and safety. Although there have been reports of severe allergic reactions caused by mRNA-LNP vaccines, the mechanism and components of anaphylaxis have not been completely clarified yet. This review focuses on two mRNA-LNP vaccines, BNT162b2 and mRNA-1273. After summarizing the structural characteristics, potential allergens, possible allergic reaction mechanism, and pharmacokinetics of mRNA and LNP in vivo, this article then reviews the evaluation methods for patients with allergic history, as well as the regulations of different countries and regions on people who should not be vaccinated, in order to promote more safe injection of vaccines. LNP has become a recognized highly customizable nucleic acid delivery vector, which not only shows its value in mRNA vaccines, but also has great potential in treating rare diseases, cancers and other broad fields in the future. At the moment when mRNA-LNP vaccines open a new era of nano medicine, it is expected to provide some inspiration for safety research in the process of research, development and evaluation of more nano delivery drugs, and promote more nano drugs successfully to market.

Objective:Mutations in the CX(connexin)26 gene (GJB2), encoding the gap\|junction protein. Connexin 26, have been shown to be the major cause of non\|syndromic recessive deafness. Among these mutations, the CX 26 cDNA (35delG) mutation accounts for the majority of this kind of deafness are European and America, but it does not present in China. Therefore, we intend to investigate the mutation hot spot of connexin 26 in China deafness populations. Mathods:The mutation of 233delC was detected by the technique of PCR\|RFLP and analyzed by directly sequencing protocols in 219 patients with kinds of deafness and 50 individuals with normal hearing. Results: The frequency of the 233delC mutation was 47/219(21.5%) in the Chinese population. No mutation was found in group with normal hearing and dominant nonsyndromic hearing loss. The mutation frequency for 233delC was 4 in 15 patients (26.7%) with genetic prelingual deafness and 10 in 50 patients(20.0%) with drug\|deafness. Conclusion: The 233delC mutaion had a high frequency in Chinese population with hearing loss, especially congenital deafness. Our data indicated that specific combinations of GJB2 mutation exist in different populations.

OBJECTIVE: To construct an adenoviral vector that codes for both human NT3 and EGFP, to confirm the transduction efficiency in rat cochlear cultures and to assess the protection of NT3 on SGNs survival. METHOD: PAdeasy-1 and pAdTrack CMV were used to constructed Ad/NT3 adenovirus and then to transfer postnatal day 3 rat cochlear cultures. The transduction efficiency was determined by microscope observation. The amounts of SGNs were counted to evaluated protection of Ad/NT3 on SGNs survival. RESULT: EGFP positive cells were observed in all cochlear turns. There was approximately 49% in outer sulcus cells and 27% in the interdental cells; less than 2% of the hair cells and SGN. The amounts of SGN of treated Ad/NT3 adenovirus are more than cochlea SGN only Ad/EGFP adenovirus after cultured for 15 days. CONCLUSION Ad/NT3 adenovirus could transduce EGFP and NT3 in large number of supporting cells, but few hair cells or SGNs. The putative release of NT3 from these supporting cells could enhance cell survival and promote neurite outgrowth from SGNs.
Adenoviridae ; genetics ; Animals ; Basilar Membrane ; cytology ; Cell Survival ; genetics ; Cells, Cultured ; Cochlea ; cytology ; Genetic Vectors ; Hair Cells, Auditory ; cytology ; Humans ; Neurotrophin 3 ; genetics ; Rats ; Rats, Inbred F344 ; Transfection

OBJECTIVE: To establish a practical model for Wistar rat cochlea organ culturing in vitro, and to observe the growing status in hypoxia of the spiral ganglion cell and nerve fiber. METHOD: We used an in vitro hypoxia model and dissociated cultures of the basal membrane from the cochlea of 3-day-old Wistar rats. And put them in incubator (37 degrees C, 90% N2, 5% CO2, 5% O2) to hypoxia culture for different times. The culture were Immunofluorescence dyed and count the number of the spiral ganglion cell and the cell density in unit area (24 mm x 36 mm), and observe the morph of nerve fiber under the confocal microscope, the results were compared with controls. RESULT: Hypoxia early (6 h) nerve fiber appear edema, spiral ganglion cell didn't change compared with controls; nerve fiber appear break and disintegration and the spiral ganglion cell decrease in 12 hours culturing, and the cell density in unit area had remarkable difference compared with control (P < 0.01). Hypoxia leads to the cell density decrease in a time-dependent manner, the longer of cultures times in hypoxia, the heavier of damage in spiral ganglion cell and nerve fiber. Twelve hours culturing, and the cell density in unit area had remarkable difference compared with control (P < 0.01). Hypoxia leads to the cell density decrease in a time-dependent manner, the longer of cultures times in hypoxia, the heavier of damage in spiral ganglion cell and nerve fiber. CONCLUSION The study findings suggest that hypoxia makes the spiral ganglion cell and nerve fiber damage of culturing in vitro, and nerve fiber more susceptible than spiral ganglion cell for hypoxia.
Animals ; Cell Count ; Cell Survival ; Female ; Hair Cells, Auditory, Inner ; cytology ; pathology ; Hypoxia ; pathology ; In Vitro Techniques ; Male ; Nerve Fibers ; pathology ; Rats ; Rats, Wistar ; Spiral Ganglion ; cytology ; pathology

OBJECTIVETo investigate the effect of Poly (ADP-ribose) polymerase (PARP) inhibition on ischemia/reperfusion (I/R) induced myocardial injury in rat and related mechanisms.

METHODAdult Wistar rats were randomly divided into sham-control (n = 18), I/R (60 min ischemia followed by 180 min reperfusion, n = 18) and I/R + PARP inhibitor 3,4-dihydro-5-[4-(1-piperidinyl)butoxy]-1(2H)-isoquinolinone (DPQ), 10 mg/kg, i.p. injection at 1 h before I/R (n = 18). Myocardial expression of PARP, infarct size, and cardiomyocytes apoptosis were determined. Additionally, myocardial NF-κB activity and the myocardial expressions of ICAM-1, COX-2 and MMP-9 at protein and mRNA level were detected.

RESULT(1) Myocardial expression of PARP was significantly upregulated in I/R group compared to sham-control group, which could be significantly reduced by pretreatment with DPQ (P < 0.05 vs. I/R group). (2) Infarct size [(31.45 ± 5.54)% vs. (45.97 ± 4.22)%] and cardiomyocytes apoptosis [(23.0 ± 3.8)% vs. (34.0 ± 6.2)%] were significantly reduced by pretreatment with DPQ (all P < 0.05 vs. I/R group). (3) Pretreatment with DPQ also significantly decreased the NF-κB activity and the myocardial expressions of ICAM-1, COX-2 and MMP-9 at both protein and mRNA level (all P < 0.05).

CONCLUSIONThe expression of PARP, NF-κB activity and the myocardial expressions of ICAM-1, COX-2 and MMP-9 are upregulated in I/R induced myocardial injury. PARP inhibitor DPQ could attenuate I/R induced myocardial injury through reducing NF-κB activity and the myocardial expressions of ICAM-1, COX-2 and MMP-9.

Animals ; Cyclooxygenase 2 ; metabolism ; Female ; Intercellular Adhesion Molecule-1 ; metabolism ; Isoquinolines ; pharmacology ; Matrix Metalloproteinase 9 ; metabolism ; Myocardial Reperfusion Injury ; drug therapy ; metabolism ; pathology ; Myocytes, Cardiac ; metabolism ; NF-kappa B ; metabolism ; Piperidines ; pharmacology ; Poly(ADP-ribose) Polymerase Inhibitors ; Poly(ADP-ribose) Polymerases ; metabolism ; Rats ; Rats, Wistar