Genomic data revealed inbreeding despite a geographically connected stable effective population size since the Holocene in the protected Formosan Long-Arm Scarab beetle, Cheirotonus formosanus.

Biodiversity conservation is a top priority in the face of global environmental change, and the practical restoration of biodiversity has emerged as a key objective. Nevertheless, the question of how to effectively contribute to biodiversity restoration and identify suitable systems for such efforts continues to present major challenges. By using genome-wide SNP data, our study revealed that populations from different mountain ranges of the Formosan Long-Arm Scarab beetle, a flagship species that receives strict protection, exhibited a single genetic cluster with no subdivision. Additionally, our result implied an association between the demographic history and historical fluctuations in climate and environmental conditions. Furthermore, we showed that, despite a stable and moderately sized effective population over recent history, all the individuals we studied exhibited signs of genetic inbreeding. We argued that the current practice of protecting the species as one evolutionarily significant unit remains the best conservation plan and that recent habitat change may have led to the pattern of significant inbreeding. We closed by emphasizing the importance of conservation genetic studies in guiding policy decisions and highlighting the potential of genomic data for identifying ideal empirical systems for genetic rescue, or assisted gene flow studies.

Testing the efficacy of different molecular tools for parasite conservation genetics: a case study using horsehair worms (Phylum: Nematomorpha).

In recent years, parasite conservation has become a globally significant issue. Because of this, there is a need for standardized methods for inferring population status and possible cryptic diversity. However, given the lack of molecular data for some groups, it is challenging to establish procedures for genetic diversity estimation. Therefore, universal tools, such as double-digest restriction-site-associated DNA sequencing (ddRADseq), could be useful when conducting conservation genetic studies on rarely studied parasites. Here, we generated a ddRADseq dataset that includes all 3 described Taiwanese horsehair worms (Phylum: Nematomorpha), possibly one of the most understudied animal groups. Additionally, we produced data for a fragment of the cytochrome c oxidase subunit I (COXI) for the said species. We used the COXI dataset in combination with previously published sequences of the same locus for inferring the effective population size (Ne) trends and possible population genetic structure.We found that a larger and geographically broader sample size combined with more sequenced loci resulted in a better estimation of changes in Ne. We were able to detect demographic changes associated with Pleistocene events in all the species. Furthermore, the ddRADseq dataset for Chordodes formosanus did not reveal a genetic structure based on geography, implying a great dispersal ability, possibly due to its hosts. We showed that different molecular tools can be used to reveal genetic structure and demographic history at different historical times and geographical scales, which can help with conservation genetic studies in rarely studied parasites.

Genetic Assessment of Fertile F1 Hybrids between Two Hercules Beetles, Dynastes maya Hardy and D. grantii Horn (Scarabaeidae).

Although hybridization may complicate taxonomic practices, it can be common between animal species. Animal hybridization not only can help with generating phenotypic and species diversity in nature, but also with understanding the genetic and genomic basis of phenotypic evolution in the laboratory. We assessed the genetic composition of captive bred F1 hybrids between two Hercules beetle species using mitochondrial CO1 and nuclear loci from a double-digest restriction-site associated DNA sequencing (ddRADseq) library. We showed that the F1 hybrids were genetically clustered with samples from the maternal species, D. grantii, based on CO1 data. Nuclear genome data, on the other hand, clearly showed that the F1 individuals were genetically intermediate between D. maya, the paternal species, and D. grantii, based on a principal component analysis. Our results also revealed that sampling design may have a major impact on the inferred genetic structure and hybrid individuals using ddRADseq data sets. We discuss the importance and potential from studying the genomics of this hybrid progeny in terms of understanding the origin and maintenance of both intraspecific and interspecific phenotypic divergence and convergence.

Intraoperative thromboelastography-guided transfusion in a patient with factor XI deficiency: A case report.

BACKGROUND: Factor XI (FXI) deficiency, also known as hemophilia C, is a rare bleeding disorder of unpredictable severity that correlates poorly with FXI coagulation activity. This often poses great challenges in perioperative hemostatic management. Thromboelastography (TEG) is a method for testing blood coagulation using a viscoelastic hemostatic assay of whole blood to assess the overall coagulation status. Here, we present the successful application of intraoperative TEG monitoring in an FXI-deficient patient as an individualized blood transfusion strategy. CASE SUMMARY: A 21-year-old male patient with FXI deficiency was scheduled to undergo reconstructive surgery for macrodactyly of the left foot under general anesthesia. To minimize his bleeding risk, he was scheduled to receive fresh frozen plasma (FFP) as an empirical prophylactic FXI replacement at a dose of 15-20 mL/kg body weight (900-1200 mL) before surgery. Subsequent FFP transfusion was to be adjusted according to surgical need. Instead, TEG assessment was used at the beginning and toward the end of his surgery. According to intraoperative TEG results, the normalization of coagulation function was achieved with an infusion of only 800 mL FFP, and blood loss was minimal. The patient showed an uneventful postoperative course and was discharged on postoperative day 8. CONCLUSION: TEG can be readily applied in the intraoperative period to individualize transfusion needs in patients with rare inherited coagulopathy.

Farnesol-Loaded Liposomes Protect the Epidermis and Dermis from PM2.5-Induced Cutaneous Injury.

Particulate matter with aerodynamic diameter ≤2.5 µm (PM2.5) increases oxidative stress through free radical generation and incomplete volatilization. In addition to affecting the respiratory system, PM2.5 causes aging- and inflammation-related damage to skin. Farnesol (Farn), a natural benzyl semiterpene, possesses anti-inflammatory, antioxidative, and antibacterial properties. However, because of its poor water solubility and cytotoxicity at high concentrations, the biomedical applications of Farn have been limited. This study examined the deleterious effects of PM2.5 on the epidermis and dermis. In addition, Farn-encapsulated liposomes (Lipo-Farn) and gelatin/HA/xanthan gel containing Lipo-Farn were prepared and applied in vivo to repair and alleviate PM2.5-induced damage and inflammation in skin. The prepared Lipo-Farn was 342 ± 90 nm in diameter with an encapsulation rate of 69%; the encapsulation significantly reduced the cytotoxicity of Farn. Lipo-Farn exhibited a slow-release rate of 35% after 192 h of incubation. The half-maximal inhibitory concentration of PM2.5 was approximately 850 µg/mL, and ≥400 µg/mL PM2.5 significantly increased IL-6 production in skin fibroblasts. Severe impairment in the epidermis and hair follicles and moderate impairment in the dermis were found in the groups treated with post-PM2.5 and continuous subcutaneous injection of PM2.5. Acute and chronic inflammation was observed in the skin in both experimental categories in vivo. Treatment with 4 mM Lipo-Farn largely repaired PM2.5-induced injury in the epidermis and dermis, restored injured hair follicles, and alleviated acute and chronic inflammation induced by PM2.5 in rat skin. In addition, treatment with 4 mM pure Farn and 2 mM Lipo-Farn exerted moderate reparative and anti-inflammatory effects on impaired skin. The findings of the current study indicate the therapeutic and protective effects of Lipo-Farn against various injuries caused by PM2.5 in the pilosebaceous units, epidermis, and dermis of skin.

Particulate matter (PM10) induces cardiovascular developmental toxicity in zebrafish embryos and larvae via the ERS, Nrf2 and Wnt pathways.

Particulate matter (PM10) is one of the most important indicators of the pollution that characterizes air quality. Epidemiological studies have shown that PM10 can cause cardiovascular-related diseases in the population. And, we studied the developmental toxicity of PM10 and the underlying mechanism of its effects on the cardiovascular system of zebrafish embryo/larva. Changes in cardiac morphology, sinus venosus and bulbus arteriosus (SV-BA) distance, heart rate, vascular subintestinalis, blood flow, returned blood volume, and reactive oxygen species (ROS) level were measured, and changes in the expression levels of certain genes were assessed via RT-PCR. The results showed that PM10 caused a significant increase in pericardial sac area and SV-BA distance, a decrease in heart rate, inhibition of vascular subintestinalis growth, blood flow obstruction, reduced venous return, and other cardiovascular toxicities. PM10 induced an increase in the ROS level and significant increases in the expression levels of ERS signalling pathway factors and Nrf2 signalling pathway factors. The expression levels of the Wnt pathway-related genes also showed significant changes. Furthermore, ROS inhibitor N-Acetyl-l-cysteine (NAC) could ameliorate the cardiovascular toxicity of PM10 in zebrafish larvae. It is speculated that PM10 may result in cardiovascular toxicity by inducing higher ROS levels in the body, which could then induce ERS and lead to defects in the expression of genes related to the Wnt signalling pathway. The Nrf2 signalling pathway was activated as a stress compensatory mechanism during the early stage of PM10-induced cardiovascular injury. However, it was insufficient to counteract the PM10-induced cardiovascular toxicity.

Isoniazid promotes the anti-inflammatory response in zebrafish associated with regulation of the PPARγ/NF-κB/AP-1 pathway.

OBJECTIVE: Zebrafish inflammation models were used to evaluate the anti-inflammatory activity of isoniazid (INH) and preliminarily investigate the underlying mechanism. METHODS: Local, acute, and systemic zebrafish inflammation models were established by tail cutting, copper sulfate (CuSO4), and lipopolysaccharide (LPS) endotoxin treatments, respectively, to evaluate the anti-inflammatory activity of INH. Zebrafish in the inflammatory state were exposed to different concentrations of INH (1, 2, and 4 mM) for 72 h to observe changes in the migration and accumulation of inflammatory cells and measure the reactive oxygen species (ROS) content in zebrafish after INH treatment. The transcription levels of inflammation-related genes in zebrafish from all groups were measured using real-time polymerase chain reaction (RT-PCR). RESULTS: Compared to those observed in the control inflammation model group, the numbers of migrated and accumulated inflammatory cells in zebrafish in the INH-treated group significantly decreased. INH significantly decreased the ROS content induced by LPS. Compared to that observed in the LPS model group, INH at 1 and 2 mM significantly increased the expression of PPARγ and inhibited the expression of NF-κB, iκbαa, and AP-1 as well as the inflammatory factors TNF-ɑ, TGF-ß, IL-1b, and COX-2. CONCLUSION: In this study, different zebrafish inflammation models were used to confirm that INH has anti-inflammatory activity. The associated mechanism may occur through the inhibition of ROS release, activation of PPARγ expression, inhibition of the transcriptional regulatory activity of NF-κB and AP-1, and reduction of INH inflammatory factor expression to relieve inflammation. The results of this study provide references for the clinical application of INH.

Effect of viscosity on pseudo-Scholte wave propagation at liquid/porous medium interface.

A theory of pseudo-Scholte wave propagating in a saturated porous medium loaded on its interface by a viscous compressible liquid is described. The porous medium is simulated by the Biot theory with high-frequency correction, and the overlying liquid is simulated by the linearized Navier-Stokes equation. An analytical expression for the complex dispersion equation of pseudo-Scholte wave through boundary conditions is established. Then the Riemann sheets related to body waves are discussed and the real and imaginary parts of the complex dispersion equation are separated and solved numerically. The resulting phase velocity, attenuation, as well as displacement and pressure fields are analyzed and comparisons are drawn with the non-viscous model. Finally, a set of parametric analyses is carried out to describe the effects of the phase velocity ratios of the S-wave in the porous medium to Ls-mode in overlying liquid on phase velocity and attenuation of the pseudo-Scholte waves.

[Establishment of An Animal Model of Transfusion-related Acute Lung Injury and the Role of Soluble CD40 Ligand in the Development of Transfusion-related Acute Lung Injury].

Objective To establish an animal model of transfusion-related acute lung injury (TRALI) and investigate the role of soluble CD40 ligand (sCD40L) in the development of TRALI. Methods The TRALI animal model established by trauma-hemorrhage-transfusion. Lung edema was evaluated by histopathological examination and the protein and Evans blue dye accumulation in bronchoalveolar lavage fluid. The concentration of sCD40L in storage packed red blood cell (PRBC) and rat's plasma was measured by enzyme-linked immunosorbent assay (ELISA). Results There were obvious epithelial hyperplasia and inflammatory cell infiltration in the lung tissue of 7 d-PRBC-treated group. The accumulation of protein in bronchoalveolar lavage fluid of 7 d-PRBC-treated group [(13.17±5.76)mg] was significantly higher than that in normal controls [(1.21±0.66)mg] and normal saline (NS)-treated group [(4.94±2.15) mg] (F=17.605,P<0.001). The leakage amount of Evans blue dye in 7 d-PRBC-treated group [(0.0109±0.0067)%/min] was significantly higher than that in NS-treated group [(0.0026±0.0006) %/min] (t=2.998,P=0.03). The concentration of sCD40L of the 7 d PRBC [(451.58±73.28) pg/ml] was significantly higher than 0 d PRBC [(277.94±98.18)pg/ml] (t=2.834,P=0.03). The concentration of sCD40L in the plasma of 7 d-PRBC-treated group [(878.21±125.30)pg/ml] was significantly higher than those in normal controls [(289.78±62.60)pg/ml] and NS-treated group [(418.07±47.68)pg/ml] (F=78.715,P<0.001). Conclusion The TRALI animal model was successfully established with trauma-hemorrhage-transfusion. The concentration of sCD40L in plasma of rats with massive transfusion is remarkably increased,suggesting sCD40L may play a role in the development of TRALI.

Comparison of learning performance of 2 intubating laryngeal mask airways in novice: A randomized crossover manikin study.

BACKGROUND: Intubating laryngeal mask airways (LMAs) such as i-gel and Aura-i could serve as rescue devices in resuscitation and further ensure the airway by facilitating trachea intubation without ventilation interruption. But data regarding intubating LMAs in novice are limited and skill degeneration without regular training has not been evaluated. So we designed this prospective randomized crossover manikin study to compare the learning performance of 2 intubating LMAs (i-gel and Aura-i). METHODS: In total, 46 novice doctors participated in this study. After standardized training and finishing 3 consecutive successful intubations with both LMAs on manikin, each participant applied intubation with both LMAs in random order for initial evaluation. To evaluate skill retention, participants were reassessed 90 days later on the same manikin without retraining between times. Primary outcome was time to successful ventilation (TTV). RESULTS: The TTV for i-gel was significantly shorter than Aura-i (initial evaluation 11.8 ±â€Š2.9 seconds vs 22.4 ±â€Š5.2 seconds, 90-days reevaluation 14.9 ±â€Š3.6 seconds vs 28.9 ±â€Š10.0 seconds, initial evaluation, P = .001; second evaluation, P < .001); during re-evaluation, TTV taken for i-gel and Aura-i were both significantly longer (initial evaluation, P = .001; second evaluation, P < .001) and ease score of insertion both increased profoundly (i-gel P = .025; Aura-i P < .001). In both assessments, participants preferred i-gel as easier alternative (initial evaluation, P = .001; second evaluation, P < .001). There was no difference in successful intubation rate, first attempt success rate, bronchoscopy assessment, and insertion score for 2 LMAs. CONCLUSION: Compared with Aura-i, i-gel showed a faster and easier intubation by novice doctors in this manikin study; the skill retention of intubation performance after 3 months was acceptable for both intubating LMAs, but TTV prolonged significantly.

Effect of Point-of-care Hemoglobin/Hematocrit Devices and Autologous Blood Salvage on Reduction of Perioperative Allogeneic Blood Transfusion.

Objective To evaluate the effect of point-of-care hemoglobin/hematocrit (POC HGB/HCT) devices and intraoperative blood salvage on the amount of perioperative allogeneic blood transfusion and blood conservation in clinical practice.Methods A total of 46 378 medical records of 22 selected hospitals were reviewed. The volume of allogeneic red blood cell and plasma, number of patients transfused, number of intraoperative autologous blood salvage, total volume of autologous blood transfusion, and amount of surgery in the year of 2011 and 2013 were tracked. Paired t-test was used in intra-group comparison, while t-test of two isolated samples carried out in inter-group comparison. P<0.05 was defined as statistically significant difference.Results In the hospitals where POC HGB/HCT device was used (n=9), the average allogeneic blood transfusion volume per 100 surgical cases in 2013 was significantly lower than that in 2011 (39.86±20.20 vs. 30.49±17.50 Units, t=3.522, P=0.008). In the hospitals without POC HGB/HCT meter, the index was not significantly different between 2013 and 2011. The average allogeneic blood transfusion volume was significantly reduced in 2013 than in 2011 in the hospitals where intraoperative autologous blood salvage ratio [autologous transfusion volume/(autologous transfusion volume+allogeneic transfusion volume)] was increased (n=12, t=2.290, P=0.042). No significant difference of the above index was found in the hospitals whose autologous transfusion ratio did not grow.Conclusion Intraoperative usage of POC HGB/HCT devices and increasing autologous transfusion ratio could reduce perioperative allogeneic blood transfusion.

Predictors for Massive Blood Loss During Posterior Correction of Congenital Scoliosis in Pre-school Children:A Retrospective Observation.

Objective To investigate the predictors for massive blood loss during posterior correction of congenital scoliosis in pre-school children. Methods Totally 124 children under six years of age,who received posterior correction of congenital scoliosis,were divided into two groups according to the ratio of intraoperative blood loss (BL) and estimated blood volume (EBV). Massive blood loss was defined as BL/EBV>0.15,and minor or moderate blood loss as BL/EBV≤0.15. All the records,including demographics,intraoperative fluids,pre- or postoperative laboratory parameters,and the length of hospital stay,were compared between these two groups. Results There were 57 children in the moderate or minor blood loss group and 67 children in the massive blood loss group. When compared with moderate or minor blood loss group,children in massive blood loss group had significantly lower body weight,shorter body height,longer anesthesia period,and more autologous or allogeneic transfusion (P<0.05). Binary Logistic regression analysis showed that body weight lower than 15 kg was the independent predictor for massive blood loss (OR=0.435,95% CI=0.197-0.962). Conclusions The incidence of massive blood loss is about 54% in children under six years of age who have received posterior correction of congenital scoliosis. The body weight of lower than 15 kg is an independent predictor for massive blood loss during the surgery.

UniQua: a universal signal processor for MS-based qualitative and quantitative proteomics applications.

Recent developments in high resolution mass spectrometry (HR-MS) technology have ushered proteomics into a new era. However, the importance of using a common, open data platform for signal processing of HR-MS spectra has not been sufficiently addressed. In this study, a MS signal processor was developed to facilitate data integration from different instruments and different proteomics approaches into a unified platform without compromising protein identification and quantitation performance. This processor supports parallel processing capability which allows full utilization of computing resources to speed up signal processing performance to >1 gigabytes/min. The storage space occupied by the processed MS data can be reduced to ~10%, which helps the analysis and management of large quantities of data from comprehensive proteomics studies. For quantitation at the MS level, processing accuracy is improved and processing time for ASAPRatio is reduced to ~50%. For quantitation at the MS/MS level, accurate reporter ion ratios from different instruments can be directly determined by the processed MS/MS spectra and reported in the Mascot search result directly without using specialized iTRAQ software.

The proto-oncogene c-src is involved in primordial follicle activation through the PI3K, PKC and MAPK signaling pathways.

BACKGROUND: C-src is an evolutionarily conserved proto-oncogene that regulates cell proliferation, differentiation and apoptosis. In our previous studies, we have reported that another proto-oncogene, c-erbB2, plays an important role in primordial follicle activation and development. We also found that c-src was expressed in mammalian ovaries, but its functions in primordial follicle activation remain unclear. The objective of this study is to investigate the role and mechanism of c-src during the growth of primordial follicles. METHODS: Ovaries from 2-day-old rats were cultured in vitro for 8 days. Three c-src-targeting and one negative control siRNA were designed and used in the present study. PCR, Western blotting and primordial follicle development were assessed for the silencing efficiency of the lentivirus c-src siRNA and its effect on primordial follicle onset. The expression of c-src mRNA and protein in primordial follicle growth were examined using the PCR method and immunohistochemical staining. Furthermore, the MAPK inhibitor PD98059, the PKC inhibitor Calphostin and the PI3K inhibitor LY294002 were used to explore the possible signaling pathways of c-src in primordial folliculogenesis. RESULTS: The results showed that Src protein was distributed in the ooplasmic membrane and the granulosa cell membrane in the primordial follicles, and c-src expression level increased with the growth of primordial follicle. The c-src -targeting lentivirus siRNAs had a silencing effect on c-src mRNA and protein expression. Eight days after transfection of rat ovaries with c-src siRNA, the GFP fluorescence in frozen ovarian sections was clearly discernible under a fluorescence microscope, and its relative expression level was 5-fold higher than that in the control group. Furthermore, the c-src-targeting lentivirus siRNAs lowered its relative expression level 1.96 times. We also found that the development of cultured primordial follicles was completely arrested after c-src siRNA knockdown of c-src expression. Furthermore, our studies demonstrated that folliculogenesis onset was inhibited by Calphostin, PD98059 or LY294002 treatment,but none of them down-regulated c-src expression. In contrast, the expression levels of p-PKC, p-ERK1/2 and p-PI3K in the follicles were clearly decreased by c-src siRNA transfection. Correspondingly, both Calphostin and LY294002 treatment resulted in a decrease in the p-PKC level in follicles, but no change was observed in the PD98059 group. Finally, LY294002 treatment decreased the p-PI3K expression level in the follicles, but no changes were observed in the PD98059 and Calphostin groups. CONCLUSIONS: C-src plays an important role in regulating primordial follicle activation and growth via the PI3K-PKC- ERK1/2 pathway.

Induction, modulation and potential targets of miR-210 in pancreatic cancer cells.

BACKGROUND: MiR-210 is induced by hypoxia and plays different roles in the development of certain cancers. However, little is known about its role in pancreatic cancer (PC). This study aimed to explore the induction and modulation of PC by miR-210 and its potential molecular targets. METHODS: PC cells were cultured under normoxic and hypoxic conditions. Expression of miR-210 and hypoxia-inducible factor (HIF)-1alpha was detected using quantitative reverse-transcription polymerase chain reaction. Cancer cells were transiently transfected with HIF-1alpha small interfering RNA (siRNA) and miR-210 mimics, and cell proliferation was measured using the CCK-8 assay. Potential targets for miR-210 were then identified using a dual luciferase reporter assay. RESULTS: Hypoxic conditions induced miR-210 expression in six PC cell lines (AsPC-1, BxPC-3, MIAPaCa-2, PANC-1, Su86.86 and SW1990), but not in Capan-1 or T3M4 cells. Transfection of HIF-1alpha siRNA into PANC-1 cells markedly inhibited HIF-1alpha expression, and subsequently down-regulated miR-210 expression under hypoxic conditions. MiR-210 had no observable impact on the proliferation of PANC-1 or Su86.86 cells and dual luciferase reporter assays showed significantly reduced luciferase activity in the wild-type E2F3, EFNA3, GIT2, MNT, ZNF462 and EGR3 constructs, compared to the corresponding mutants, but not in HOXA3. CONCLUSIONS: These results suggest that miR-210 expression in PC cells is induced by hypoxia through a HIF-1alpha-dependent pathway, but does not influence PC cell proliferation. Also, E2F3, EFNA3, GIT2, MNT, ZNF462 and EGR3 may be potential miR-210 targets in PC.

Circulating microRNAs in cancer: diagnostic and prognostic significance.

The involvement of microRNAs (miRs) in numerous pathological conditions is well established. In many kinds of cancer cells and animal models, various miRs have been shown to act as oncogenes or tumor suppressors. Recently, it was found that circulating miRs can be detected, and may be associated, with the clinicopathological features and prognosis of cancers, thus, providing potential novel diagnostic and prognostic markers for malignancies in humans. This review aims to address these issues based on recently published literature.

Tunnel frit: a nonmetallic in-capillary frit for nanoflow ultra high-performance liquid chromatography-mass spectrometryapplications.

In this study, an easy method to fabricate a durable in-capillary frit was developed for use in nanoflow liquid chromatography (nanoLC). A small orifice was tunneled into the sol-gel frit during the polymerization process resulting in the simple fabrication of a tunnel frit. A short packing tunnel frit column (2 cm, C(18) particles) was able to sustain over 10,000 psi continuous liquid flow for 10 days without observation of particle loss, and back pressure variation was less than 5%. The tunnel frit was successfully applied to the fabrication of nanoflow ultra high-performance liquid chromatography (nano-UHPLC) trap and analytical columns. In the analysis of tryptic peptides, the tunnel frit trap and analytical columns were demonstrated to have high separation efficiency and sensitivity. In analysis of phosphopeptides, the use of the nonmetallic tunnel frit column showed better sensitivity than the metallic frit column. This design can facilitate the preparation of nano-HPLC and nano-UHPLC columns and the packing material can easily be refilled when the column is severely contaminated or clogged.

[Preliminary study about the role of c-src in the initiation of primordial follicle in rat ovary].

OBJECTIVE: To explore the role of c-src on the initiation of primordial follicles. METHODS: 2-days-old female SD rats' ovaries were cultured in Waymouth culture system and were used HE staining and immunohistochemy to observe the number of follicles after 0, 4, 8 days cultured. Use chemically synthesized small interference RNA (siRNA) transfected into ovarian tissue in cultured for RNA interference, and use HE staining and RT-PCR to detect the best siRNA and packaging it by lentiviruses to test the interference effect. RESULTS: With the increase of culturing days, the nummber of the primordial follicles in ovarian gradually reduced. We packed the best siRNA by lentiviruses to doing RNA interference and found comparing with the blank control group and blank vector group, c-src mRNA of the best interference group were significantly decreased. The total number of primordial follicles was relatively greater and the development of primordial folliculars was inhibited. CONCLUSION: c-src plays an important role in primordial follicle development and folliculogenesis initiation.

[The expression of proto-oncogene c-erbB2 and its role in the initiation of primordial follicle growth in rat ovary].

Little is known about the factors that control the initiation of growth of primordial follicles. The objective of the present study was to investigate the effect of c-erbB2 on the onset of primordial follicle development, and whether c-erbB2 mediates the effect of epidermal growth factor (EGF) in this process. We synthesized three pairs of siRNAs targeting the c-erbB2 mRNA and transferred them into the newborn rat ovary cultured in vitro with Metafectene. After siRNAs transfection, the efficiency of siRNAs was tested by examining c-erbB2 mRNA and protein levels. The level of c-erbB2 mRNA was reduced by 49.6%, 46.7% and 82.6% respectively after transfecting siRNA1, siRNA2 and siRNA3, and the level of ErbB2 protein also reduced remarkably after siRNA3 transfection. c-erbB2 siRNA3 significantly inhibited the primordial follicle initiation and development; EGF augmented primordial follicles formation, but the effect was abolished by c-erbB2 siRNA3. All of these results suggest that c-erbB2 plays an important role in primordial follicle development and folliculogenesis initiation, and mediates the effect of EGF on primordial follicle development.

Molecular identification of forensically important blow fly species (Diptera: Calliphoridae) in Taiwan.

Forensic entomology is a discipline that mainly uses insects collected in and around corpses to estimate the post-mortem interval in medicocriminal investigations. Among all scavenger and necrophagous insect groups that are related to corpses, blow flies (Diptera: Calliphoridae) are probably most important, not only because they occur in abundant numbers but also because they are one of the earliest groups to find corpses. However, most entomological evidence is strongly dependent on accurate species identification. Because identification allows the proper developmental data and distribution ranges to be applied in criminal investigations, species in Taiwan were surveyed from early 2000 and were identified using molecular data. Currently, eight species have been identified: Chrysomya megacephala (Fabricius), Chrysomya pinguis (Walker), Chrysomya rufifacies (Macquart), Hemipyrellia ligurriens (Wiedemann), Lucilia bazini Séguy, Lucilia cuprina (Wiedemann), Lucilia hainanensis Fan, and Lucilia prophyrina (Walker). We focused on classifying these blow fly species to establish a knowledge basis for further forensic entomological research in Taiwan. Because molecular data are helpful in identifying insect specimens, especially when no specimen of suitable condition for morphological identification is obtained, we extracted mitochondrial cytochrome oxidase subunit I (COI) DNA of the preceding blow fly species to study its application value for their differentiation. The cloning and sequencing of the COI gene (approximately 1,588 base pairs) of these eight species were completed, and the data were analyzed. Preliminary results revealed the high support of congeneric groupings of species by using COI data; these sequences were also shown to be highly conserved within the same species. To actually use the database of COI sequences under various specimen conditions, specific primers were also applied for different insect stages, different segments of adults, and specimens preserved for various times. A molecular primer key was ultimately constructed for the purpose of rapid and accurate species identification at the molecular level regardless of which stage or which part of a blow fly specimen is collected.