[Application of near infrared fluorescence imaging in detection of residual cancer in oral squamous cell carcinoma].

Objective: Local recurrence is the main cause of treatment failure in patients with oral squamous cell carcinoma (OSCC). This study was proposed to investigate the feasibility of near infrared fluorescence (NIF) via indocyanine green (ICG) for monitoring surgical marginal in operation for OSCC patients. Methods: In 35 patients with OSCC treated surgically in the Department of Oral and Maxillofacial Surgery, Nanjing University School of Medicine, from January 2019 to June 2020, ICG (0.75 mg/kg) was administered intravenously via elbow vein at (12±1) hours before surgery, and NIF was performed intraoperatively on the surgical field and the cut edge of the surgically excised specimen, and fluorescence intensity was measured for OSCC tissue and normal oral mucosa, abnormal fluorescence signals were taken and subjected to rapid cryopathological examination. Correlation between NIF tumor boundary grading and pathological tumor boundary grading was analyzed by Spearman correlation analysis. Results: Clear ICG NIF was obtained for tumor lesions in all 35 patients, with a positive rate of 100%. The fluorescence intensity of OSCC tissue was (412.73±146.56) au, which was higher than that of normal oral mucosa tissue [(279.38±82.56) au, P<0.01]. Abnormal fluorescence signals were detected at the tumor bed and the cut edge of the surgical resection specimen in 4 patients, of which 2 cases were pathologically confirmed as cancer cell residue and 2 cases as inflammatory cell infiltration. The rate of positive detection of cut margins using ICG NIF technique in OSCC was 5.7% (2/35). Twenty of the 35 OSCC patients had grade 1, 11 of grade 2, and 4 of grade 3 tumor borders revealed by NIF of surgical resection specimens, which was positively correlated with pathological tumor border (r=0.809, P<0.001). Conclusions: ICG NIF technique can effectively detect the residual cancer cells at the incision margin, which is of great clinical value in reducing local recurrence of OSCC after surgery due to intraoperative cancer residue.

Effects of nonglycosylated and glycosylated prolactin on basal and gonadotropin-stimulated steroidogenesis in chicken ovarian follicles.

In galliformes, the circulating isoform of prolactin (PRL) significantly changes during different reproductive states. However, the role of the major isoform (glycosylated PRL [G-PRL]) in ovarian steroidogenesis is unknown. The present study aimed to compare the effects of nonglycosylated (NG-) and G-PRL on basal and gonadotropin-stimulated estradiol (E2) and progesterone (P4) production in granulosa cells or follicular walls of chicken of different size class follicles. In the initial experiment, granulosa cells of preovulatory F3-F1 and prehierarchical 6- to 8-mm follicles were incubated for 24 h with different concentrations of NG- or G-PRL (0, 1, 10, 100, or 1,000 ng/mL). In the subsequent experiments, these categorized granulosa cells and follicular walls of prehierarchical 4-6, 2-4, and <2-mm follicles were incubated for 24 h in the absence and presence of 10-ng/mL FSH or LH, or in combination with different concentrations of NG- or G-PRL (10, 100, or 1,000 ng/mL). We observed that lower levels of NG-PRL induced (P < 0.05) E2 and P4 secretion in granulosa cells of either preovulatory or prehierarchical follicles, but at higher levels, this effect was reduced. In contrast, G-PRL promoted (P < 0.05) basal E2 and P4 secretion in preovulatory granulosa cells but was inhibitory (P < 0.05) in prehierarchical granulosa cells. Results obtained by real-time quantitative PCR (qPCR) demonstrated that these effects were mediated through modulation of the expression of StAR, CYP11A1, CYP19A1, and 3ß-HSD. Furthermore, G-PRL was less potent than NG-PRL in inhibiting FSH- or LH-stimulated E2 and P4 production in granulosa cells of preovulatory follicles, whereas NG-PRL enhanced (P < 0.05) but G-PRL reduced (P < 0.05) FSH-induced P4 production in those of prehierarchical follicles. In follicular walls from each group of prehierarchical 4-6, 2-4, and <2-mm follicles, NG- and G-PRL had both stimulatory and inhibitory influences on the actions of FSH on E2 and P4 secretion, but both suppressed (P < 0.05) LH-induced E2 and P4 secretion except for the synergistic effects of LH and G-PRL on P4 secretion by follicular walls of the follicles of 4-6 mm. Taken together, these results suggest that both NG- and G-PRL are biologically active in regulating basal and gonadotropin-stimulated E2 and P4 production in chicken ovarian follicles. However, their effects are different depending on the concentration, the type of gonadotropin (FSH or LH), and the stage of follicle development.

The renin-angiotensin system blockers as adjunctive therapy for cancer: a meta-analysis of survival outcome.

OBJECTIVE: The renin-angiotensin system blockers (RASBs), including angiotensin-converting enzyme inhibitors (ACEIs) and angiotensin II receptor blockers (ARBs), are widely used to reduce cardiovascular disease (CVD) events. Accumulating pre-clinical and clinical studies suggested that RASBs possesses anti-carcinogenic properties, and their use is associated with favorable outcomes in several type cancers. To conduct a meta-analysis to evaluate the effect of adjunctive therapy of renin-angiotensin system blockers combined with chemotherapeutic agents in cancer patients. MATERIALS AND METHODS: Data from a total of 2436 patients from 7 retrospective studies investigating chemotherapeutic agents in combination with RASBs agents versus chemotherapeutic agents were included in this meta-analysis. Publication bias was assessed by the Begg's Test, Egger's test and funnel plot. Subgroup analysis was conducted when the chemotherapeutic agents were the same. RESULTS: A significant reduction in overall mortality in favor of chemotherapeutic agents in combination with RASBs agents was observed, hazard ratio (HR) 0.80 (95% CI: 0.69-0.92); there was a significant decrease in the risk of disease progression in favor of chemotherapeutic agents in combination with RASBs regimens, HR 0.79 (95% CI: 0.66-0.94), compared with those who only used chemotherapeutic agents. Subgroup analysis indicated that platinum-based agents plus ACEI/ARB could increase significantly the survival outcome (HR = 0.56; 95% CI: 0.38-0.82). CONCLUSIONS: Our results suggest that RASBs combined with chemotherapeutic agents may improve outcomes in multiple types' cancer patients. More research and well-designed, rigorous, large clinical trials are required to address these issues.

[The clinical features of patients with allergic rhinitis of Nantong city in China].

Objective:To comprehensively analyze the clinical features of patients with AR by a retrospective study. Method:A total number of 8 102 patients diagnosed with AR were enrolled in Nantong area, and detailed clinical data were documented in all cases. Skin prick tests with standardized aeroallergens were conducted in these patients; The samples were divided into two groups(children and adults group) and the clinical features between two groups were analysed. Result:Children in schoolage were the majority of AR patients in children group. There were 4 581 cases(56.54%) with ocular symptoms, 3 977 cases(49.09%)with lower respiratory tract symptoms. Nasal congestion (97.37%) was the most common symptoms in patients with AR,while eye itching(32.68%) was the most common ocular symptoms in patients with AR, followed by the dacryorrhea(23.57%);and cough(44.72%) was the most common lower respiratory tract symptoms. Dermatophagoides pteronyssinus and dermatophagoides farinae had the highest positivity among all allergens, and the shrimp was the main food allergen. Conclusion:We analyzed the clinical features of patients with AR, that would provide a more scientific basis for prevention,clinical diagnosis, treatment and epidemiological studies for AR.

Applicability of first-trimester combined screening for fetal trisomy 21 in a resource-limited setting in mainland China.

OBJECTIVE: To assess the feasibility and performance of the first-trimester combined screening test for trisomy 21 in a resource-limited setting in mainland China. DESIGN: Prospective observational cohort study. SETTING: First Affiliated Hospital of Kunming Medical University, China. POPULATION: Ten thousand four hundred and forty-two pregnant women requesting first-trimester screening. METHODS: The combined screening test was performed from May 2012 to December 2014. Women with a high-risk result (≥1:600) were offered further confirmatory tests after counselling. The threshold for high risk was determined by Monte Carlo simulation to achieve a 5% false-positive rate according to the local age distribution. Pregnancy outcome and screening results were recorded for all women and monthly audits were conducted. MAIN OUTCOME MEASURES: Sensitivity, screen positive rate, cost per case of Down syndrome detected. RESULTS: Six hundred and ten women (5.8% of the total screened) had a high-risk screening test, of whom 274 (44.9%) underwent a diagnostic test and 169 (27.7%) opted for a noninvasive prenatal screening test (NIPT); 160 (26.2%) declined further testing after counselling. The pregnancy outcome was available for 10 174 (97.4%) of the women. The observed incidence of Down syndrome was 0.13% (1/750). All 14 women with a trisomy 21 pregnancy had a high-risk screening test result. The cost per Down syndrome detected was RMB596 686 compared with RMB1.79 million if all had been screened by NIPT. CONCLUSIONS: The combined screening test appears to be a more cost-effective strategy in mainland China. Screening performance in China would be improved by adopting Chinese-specific models, external quality control and assurance, and establishing risk thresholds appropriate for the age distribution of the population. TWEETABLE ABSTRACT: Combined first-trimester Downs screening in China was improved by adopting Chinese-specific models and external QC.

Losartan improves the distribution and efficacy of doxorubicin in CT26 tumor.

OBJECTIVE: The effectiveness of chemotherapeutic agents is impaired by limited delivery of chemotherapeutic agents to the tumor cells. Improving drug penetration in tumor tissues is very important. We tested whether losartan, a selective antagonist against type 1 angiotensin II receptors (AT1R) with noted antifibrotic activity, can enhance the penetration and efficacy of doxorubicin. MATERIALS AND METHODS: BALB/C mice, which implanted with CT26 tumor cells, were divided into four groups: control, doxorubicin alone, losartan alone and doxorubicin + losartan combination groups. At day 0, the losartan alone and doxorubicin + losartan combination groups received losartan; and at day 8, the doxorubicin alone and doxorubicin + losartan combination groups received doxorubicin i.v. Tumor growth and intratumoral distribution of doxorubicin were evaluated. The mechanism underlying the enhanced anti-tumor effect of the combination of doxorubicin and losartan was investigated by immunohistochemical analysis. RESULTS: Treatment with losartan alone did not suppress tumor growth; In contrast, treatment with doxorubicin alone decreased tumor growth; losartan and doxorubicin were administered in combination, had a synergistic effect that the tumor growth was much more inhibited. The decreased proliferation as indicated by down-regulation of Ki67, and increased apoptosis as indicated by TUNEL and caspase-3 staining. The expression of tumor suppressor gene P53 increased in doxorubicin + losartan combination groups. CONCLUSIONS: Losartan can increase the therapeutic effectiveness of doxorubicin, yielding more great antitumor benefit. This study provided a rationale for initiating clinical trials using losartan in combination with chemotherapeutic agents to increase their therapeutic effectiveness.

The cancer-associated fibroblasts and drug resistance.

The present treatment of solid tumors is plagued by drug resistance. Despite continued development of meticulously designed therapeutic scheme, cancer cells remain poorly being completely eliminated. Because therapeutic resistance is a problem with the drug used in cancer therapy, most of the studies about the drugs resistance have focused on the role of epithelial cancer cells themselves. However, it is becoming increasingly apparent that tumor microenvironment could provide a shelter for tumor cells which keep their survival after initial drug exposure. Cancer-associated fibroblasts (CAFs) are the crucial component of the tumor microenvironment; substantial evidence suggests that the CAFs mediate resistance of solid tumor cells to the anticancer drugs. In this review, we describe how the CAFs may be involved in the resistance of tumor cells to the therapeutic agents and present some of the emerging therapeutic targets for modulation this resistant phenotype.

Endogenous nitric oxide induces activation of apoptosis signal-regulating kinase 1 via S-nitrosylation in rat hippocampus during cerebral ischemia-reperfusion.

Apoptosis signal-regulating kinase 1 (ASK1) is a general mediator of cell death in response to a variety of stimuli, including reactive oxygen species, tumor necrosis factor α, lipopolysaccharide, endoplasmic reticulum stress, calcium influx and ischemia. Here we reported ASK1 was activated by nitric oxide (NO) through S-nitrosylation during cerebral ischemia-reperfusion. The reagents that abrogate neuronal nitric oxide synthase (nNOS) activity such as nNOS inhibitor 7NI and N-methyl-D-aspartate receptor antagonist MK801 prevented ASK1 activation via decreasing ASK1 S-nitrosylation. In HEK293 cells, over-expressed ASK1 could be S-nitrosylated by both exogenous and endogenous NO and Cys869 was identified as the site of ASK1 S-nitrosylation. S-nitrosylation increased the level of ASK1 phosphorylation at Thr845, which represents ASK1 activation. Our results further confirmed that S-nitrosylation led to the increment of ASK1 dimerization. S-nitrosylation of ASK1 also activated the downstream JNK signaling and JNK-mediated nucleic pathway. The exogenous NO (SNP and GSNO) reversed the effect of endogenous NO by suppressing S-nitrosylation of ASK1 and exerted neuroprotection during ischemia-reperfusion. These results suggest that inhibiting ASK1 S-nitrosylation may be a novel approach for stroke therapy.

Hierarchical protein folding: asymmetric unfolding of an insulin analogue lacking the A7-B7 interchain disulfide bridge.

The landscape paradigm of protein folding can enable preferred pathways on a funnel-like energy surface. Hierarchical preferences may be manifest as a nonrandom pathway of disulfide pairing. Stepwise stabilization of structural subdomains among on-pathway intermediates is proposed to underlie the disulfide pathway of proinsulin and related molecules. Here, effects of pairwise serine substitution of insulin's exposed interchain disulfide bridge (Cys(A7)-Cys(B7)) are characterized as a model of a late intermediate. Untethering cystine A7-B7 in an engineered monomer causes significantly more marked decreases in the thermodynamic stability and extent of folding than occur on pairwise substitution of internal cystine A6-A11 [Weiss, M. A., Hua, Q. X., Jia, W., Chu, Y. C., Wang, R. Y., and Katsoyannis, P. G. (2000) Biochemistry 39, 15429-15440]. Although substantially disordered and without significant biological activity, the untethered analogue contains a molten subdomain comprising cystine A20-B19 and a native-like cluster of hydrophobic side chains. Remarkably, A and B chains make unequal contributions to this folded moiety; the B chain retains native-like supersecondary structure, whereas the A chain is largely disordered. These observations suggest that the B subdomain provides a template to guide folding of the A chain. Stepwise organization of insulin-like molecules supports a hierarchic view of protein folding.

[Expression and purification of murine interleukin 18 in Escherichia coli and its antitumor effects].

Total RNA was extracted from murine hepatocytes, and the cDNA of interleukin 18(IL-18) was amplified by RT-PCR. The cDNA was introduced into the expression vector pJW2 and sequenced. Under heat induction, the recombinant murine IL-18(rmIL-18) was expressed in inclusion bodies in E. coli with the yield accounting for 18% of total bacteria proteins. The inclusion bodies were dissolved with 5 mol/L urea, and rmIL-18 was purified using Sephadex G-100 column chromatography. In the presence of 0.5 mg/L Con A, the purified rmIL-18 showed dose-dependent IFN-gamma-inducing activity in murine splenocytes. The purified rmIL-18 exhibited significant antitumor effects in Kunming mice challenged intraperitoneally (i.p.) with H22 hepatocarcinoma when administered 10 micrograms rmIL-18 i.p. on days 1, 4 after challenge, and the mice survived resisted the rechallenged with H22 cells.

Mapping the functional surface of insulin by design: structure and function of a novel A-chain analogue.

Functional surfaces of a protein are often mapped by combination of X-ray crystallography and mutagenesis. Such studies of insulin have yielded paradoxical results, suggesting that the native state is inactive and reorganizes on receptor binding. Of particular interest is the N-terminal alpha-helix of the A-chain. Does this segment function as an alpha-helix or reorganize as recently proposed in a prohormone-convertase complex? To correlate structure and function, we describe a mapping strategy based on protein design. The solution structure of an engineered monomer ([AspB10, LysB28, ProB29]-human insulin) is determined at neutral pH as a template for synthesis of a novel A-chain analogue. Designed by analogy to a protein-folding intermediate, the analogue lacks the A6-A11 disulphide bridge; the cysteine residues are replaced by serine. Its solution structure is remarkable for segmental unfolding of the N-terminal A-chain alpha-helix (A1 to A8) in an otherwise native subdomain. The structure demonstrates that the overall orientation of the A and B chains is consistent with reorganization of the A-chain's N-terminal segment. Nevertheless, the analogue's low biological activity suggests that this segment, a site of clinical mutation causing diabetes mellitus, functions as a preformed recognition alpha-helix.

Insulin-like compounds related to the amphioxus insulin-like peptide.

Three insulin-like compounds consisting of two disulfide-linked polypeptide chains have been synthesized. The A-chains of these compounds correspond either to the A- or to the A + D-domain of the putative amphioxus insulin-like peptide (amphioxus ILP), and their B-chains correspond either to the B-chain of insulin or to a slightly modified (i.e., [1-Thr]) B-domain of amphioxus ILP. The biological potency of these compounds was evaluated in mammalian cells or cell fractions containing either human or rat insulin receptors or human or mouse insulin-like growth factor I (IGF-I) receptors, with respect to binding affinity, insulin-like metabolic activity (lipogenesis), and growth factor activity (mitogenesis). Amphioxus ILP A/bovine insulin B and amphioxus ILP A + D/bovine insulin B exhibited potencies ranging from 2.0 to 9.8% relative to natural insulin, and both compounds were full agonists in lipogenesis assays, stimulating lipogenesis to the same maximal extent as seen with natural insulin. Amphioxus ILP A/amphioxus ILP [1-Thr]B stimulated lipogenesis with a potency of 0.01% relative to natural insulin. We consider this compound also likely to be a full agonist. In assays measuring binding to IGF-I receptors and stimulation of mitogenesis, these compounds displayed some activity although the activity was too low for exact quantification. These results suggest that amphioxus ILP has retained an overall structural similarity to mammalian insulin and IGF-I but has also accumulated substantial mutations which markedly reduce its ability to bind and activate their cognate receptors.(ABSTRACT TRUNCATED AT 250 WORDS)

[Changes in water-soluble, urea-soluble and membrane intrinsic proteins in human senile cataract].

Gel filtration of water-soluble protein shows a substantial increase in HM+ alpha-crystallin and a marked decrease in beta- and gamma-crystallins in cortical cataract. A decrease in beta 1-crystallin in cortical punctate opaque lenses is also striking. In nuclear cataractous lenses HM+ alpha- and beta-crystallin increase, while gamma-crystallin decreases. The urea-soluble protein from clear lenses contains mainly of alpha beta chain, whereas in cataractous lenses the relative amounts of the 28 and 23ku polypeptides (the components of beta-crystallin) increased markedly. In cataractous lenses the relative amount of membrane intrinsic proteins decreases slightly and it has little statistical meaning.