Transcripts expressed using a bicistronic vector pIREShyg2 are sensitized to nonsense-mediated mRNA decay

<p>Abstract</p> <p>Background</p> <p>pIREShyg2 has been widely used as a bicistronic expression vector. However, it is not known if the vector would affect the expression of cloned genes via nonsense-mediated mRNA decay (NMD), an mRNA surveillance system that degrades mRNA with a premature termination codon (PTC). In mammalian cells, the induction of NMD requires either a long 3'UTR or the presence of an exon-junction complex downstream of a PTC. The efficiency of NMD is greater when a PTC generates longer 3'UTR. pIREShyg2 provides the first cistron gene with a long 3'UTR consisting of a downstream intervening sequence (IVS), an internal ribosomal entry site (IRES) and the second cistron. Therefore, we hypothesized that the first cistron genes in pIREShyg2 are sensitized to NMD, which affects their expression levels. To examine this hypothesis, cDNAs encoding human granulocyte-macrophage colony-stimulating factor receptor β chain (βc) and its splice variant (βc79), in which the retention of a 79-base intron caused a frameshift generating 18 PTCs, were cloned into pIREShyg2 and stably expressed in a murine cell line, Ba/F3.</p> <p>Results</p> <p>Compared with wild-type βc, the mRNA levels of βc79 were less than one tenth and decayed faster. Both translation inhibition and Upf1 knockdown led to significantly greater up-regulation of βc79 than wild-type βc. However, the use of a monocistronic pMT21 vector abolished the up-regulatory effects of translation inhibition and Upf1 knockdown on both wild-type βc and βc79, suggesting that the NMD is attributable to a structural determinant in pIREShyg2. The elimination of the intron and the proximal 3' 17 PTCs did not alter the greater effects of translation inhibition on βc79, suggesting that the first PTC, which determines 3'UTR length, was sufficient to enhance NMD efficiency. Thus, transcripts of PTC-harboring genes with longer 3'UTR are more efficiently degraded by the vector-dependent NMD than those of wild-type genes with relatively shorter 3'UTR, resulting in minimized expression of truncated mutants.</p> <p>Conclusions</p> <p>We conclude that pIREShyg2, which sensitizes its bicistronic transcripts to NMD, may be useful for studying NMD but should be avoided when maximum expressions of PTC-harboring genes are required.</p

COMBINATION THERAPY OF INTRAVITREAL RANIBIZUMAB AND SUBTHRESHOLD MICROPULSE PHOTOCOAGULATION FOR MACULAR EDEMA SECONDARY TO BRANCH RETINAL VEIN OCCLUSION

Purpose: To determine the efficacy of the combination therapy of intravitreal ranibizumab (IVR) and 577-nm yellow laser subthreshold micropulse laser photocoagulation (SMLP) for macular edema secondary to branch retinal vein occlusion cystoid macular edema.Methods: Retrospective, consecutive, case–control study. Forty-six eyes of 46 patients with treatment-naive branch retinal vein occlusion cystoid macular edema were enrolled. The IVR + SMLP group consisted of 22 patients who had undergone both SMLP and IVR. Intravitreal ranibizumab group consisted of 24 patients who had undergone IVR monotherapy. Intravitreal ranibizumab therapy was one initial injection and on a pro re nata in both groups, and SMLP was performed at 1 month after IVR in the IVR + SMLP group. Preoperatively and monthly, best-corrected visual acuity and central retinal thickness were evaluated using swept source optical coherence tomography.Results: Best-corrected visual acuity and central retinal thickness significantly improved at 6 months in IVR + SMLP and IVR groups. Best-corrected visual acuity and central retinal thickness were not significantly different between the two groups at any time points. The number of IVR injections during initial 6 months in IVR group (2.3 ± 0.9) was significantly greater (P = 0.034) than that in IVR + SMLP group (1.9 ± 0.8).Conclusion: The combination therapy of IVR and SMLP can treat branch retinal vein occlusion cystoid macular edema effectively, by decreasing the frequency of IVR injections while maintaining good visual acuity

Effects of a novel method of anesthesia combining propofol and volatile anesthesia on the incidence of postoperative nausea and vomiting in patients undergoing laparoscopic gynecological surgery

AbstractBackgroundWe investigated the effects of a novel method of anesthesia combining propofol and volatile anesthesia on the incidence of postoperative nausea and vomiting in patients undergoing laparoscopic gynecological surgery.MethodsPatients were randomly divided into three groups: those maintained with sevoflurane (Group S, n=42), propofol (Group P, n=42), or combined propofol and sevoflurane (Group PS, n=42). We assessed complete response (no postoperative nausea and vomiting and no rescue antiemetic use), incidence of nausea and vomiting, nausea severity score, vomiting frequency, rescue antiemetic use, and postoperative pain at 2 and 24h after surgery.ResultsThe number of patients who exhibited a complete response was greater in Groups P and PS than in Group S at 0–2h (74%, 76% and 43%, respectively, p=0.001) and 0–24h (71%, 76% and 38%, respectively, p<0.0005). The incidence of nausea at 0–2h (Group S=57%, Group P=26% and Group PS=21%, p=0.001) and 0–24h (Group S=62%, Group P=29% and Group PS=21%, p<0.0005) was also significantly different among groups. However, there were no significant differences among groups in the incidence or frequency of vomiting or rescue antiemetic use at 0–24h.ConclusionCombined propofol and volatile anesthesia during laparoscopic gynecological surgery effectively decreases the incidence of postoperative nausea. We term this novel method of anesthesia “combined intravenous-volatile anesthesia (CIVA)”

Plasma S100A12 Levels and Peripheral Arterial Disease in End-Stage Renal Disease

Background: S100A12 is an endogenous ligand of the receptor for advanced glycation end products (RAGE). Plasma S100A12 levels are high in end-stage renal disease (ESRD) patients undergoing maintenance hemodialysis (HD). Peripheral arterial disease (PAD) is common in HD patients and is associated with increased cardiovascular morbidity and mortality rates in this population. To date, however, no study has specifically assessed the relationship between plasma S100A12 and PAD in HD patients. Methods: We conducted a cross-sectional study of 152 HD patients in our affiliated hospital. We investigated PAD history and patient characteristics and quantified plasma S100A12 levels in all participants. Results: HD patients with PAD (n = 26; 21.9 [13.6–33.4] ng/ml) showed significantly higher plasma S100A12 levels than HD patients without PAD (n = 126; 11.8 [7.5–17.6]ng/ml; p Conclusion: These results suggest that plasma S100A12 levels are strongly associated with PAD prevalence in ESRD patients undergoing HD

Heating Has No Effect on the Net Protein Utilisation from Egg Whites in Rats

Egg whites (EW) are a good source of protein; however, they are typically heated prior to consumption. Therefore, we investigated the effects of different heating conditions on the protein utilisation rate of EW. Male Sprague-Dawley rats (n=36, 198±1 g) were divided into six groups and fed American Institute of Nutrition-76 chow containing unheated EW, soft-boiled EW, boiled EW, milk whey protein, soybean protein, or no protein over a 10-day period using pair-feeding. Urine and faeces were sampled daily beginning on day 5 to measure nitrogen content and the net protein utilisation (NPU) rate. The soybean protein group had a significantly lower level of food intake and was thus excluded from subsequent analyses. The NPU value was similar among the unheated, soft-boiled, and boiled EW groups (97.5±0.4, 96.5±0.1, and 96.5±0.7, resp.). The EW group values were significantly higher than the whey group values (90.5±1.0). These results show that EW serve as a good source of protein, irrespective of heating

Successful pregnancy with stage IB2 uterine cervical cancer: A case report

Abstract Background Although cervical cancer is one of the most common malignancies in pregnancy, its management mainly follows the guidelines for nonpregnant disease state. Within the limited time, patients, and healthcare workers must make difficult decisions to either delay treatment until documented fetal maturity or start immediate treatment based on the disease stage. Case The patient was a 37‐year‐old woman: gravida 1, para 0. Her cervical cytology revealed a high‐grade squamous intraepithelial lesion at 8 weeks' gestation. Moreover, invasive squamous cell carcinoma was suspected based on the findings of uterine cervix biopsy. Cervical conization was performed at 11 weeks' gestation, confirming a histopathological diagnosis of squamous cell carcinoma, pT1b2. Cervical cytology findings continued to be negative for intraepithelial lesion or malignancy from 2 weeks after conization until 2 weeks before a cesarean section. In addition, we performed abdominal pelvic lymphadenectomy at 16 weeks' gestation to determine whether the patient could continue her pregnancy. No lymph node metastasis or local recurrence was observed. Finally, a cesarean section and modified radical hysterectomy were performed at 35 weeks' gestation. There was no carcinoma invasion or metastasis. A baby girl weighing 2056 g was delivered with 1‐ and 5‐min Apgar scores of 8 and 9, respectively. Five years postoperatively, there was no evidence of cancer recurrence. Conclusion Management of cervical cancer during pregnancy by using a combination strategy of deep conization and pelvic lymphadenectomy could be an effective strategy for carefully and safely assessing risks of recurrence and metastasis