Structural Aberrations of Cellular Sialic Acids and Their Functions in Cancer Metastases

Sialic acids (neuraminic acids) are a special series of 9 carbon ring negatively charged carbohydrates, which has been found to be selectively changed in malignant cells from structures (both synthesis and structure modifications) to functions (up and down regulation in cells). Sialic acids, in single forms or conjugates, have been systematically studied both in lab and in clinics by GC, GC MS, NMR, HPTLC, HPLC and other modern analytical means. Sialic acids and related conjugates are predicted to be used in cancer diagnosis, cancer prognostic forecasting, designing of cancer chemotherapy regimens, uncovering carcinogenetic processes and neoplasm metastasis. Tumor cell regulative systems and pathways are correlated with sialic acids, which can be applied to prognostic evaluation of cancer patients, and antimetastatic chemotherapy by sialic acid derivatives and analogues. Searching for new biological characteristics of sialic acids in cells have also been extensively studied these days. In this paper, main stream discoveries and advancements are provided , also discussions of possible mechanisms and hypotheses are invoked.

Blockade of neutrophil recruitment to tumor sites based on sialic acid-modified nanoplatforms enhances the efficacy of checkpoint blockade immunotherapy

Checkpoint inhibitors are designed to rejuvenate depleted or suppressed T cells in the tumor microenvironment,relying on the immune system to control and kill tumors.However,accumulating evidence indicates that tumor-infiltrating neutrophils impede the proliferation and activation of T cells and determine the resistance to checkpoint blockade and chemotherapy.In this study,sialic acid ligand-modified colchicine derivative phospholipid complexes specifically targeted tumor-associated neutrophils in the peripheral blood,blocked neutrophil accumulation in tumors,and attenuated the inhibitory effect of infiltrating neutrophils on T cells.Neutrophil blocking therapy enhanced the immunotherapy effect of the PD-L1 antibody in S180 advanced tumors and 4T1 breast cancer.Our study found that PD-L1 antibody monotherapy increased the tumor infiltration of immunosuppressive neutrophils.Combination therapy with neutrophil blocking can greatly reduce tumor-infiltrating neutrophils and increase the proliferation of cytotoxic CD8^(+) T lymphocytes in the tumor.The combination therapy significantly improved the survival rate of mice with advanced S180 tumors and increased the sensitivity of immune checkpoint inhibitors to 4T1 cold tumors.

New Extracellular Bacillus subtilis Lectins with Sialic Acid-Specificity

The purpose of this work is to perform a detailed study of carbohydrate specificity of the new extracellular bacilli lectins which is considered to determine mechanisms of the lectins action. Sources of lectins were bacterial strains from Ukrainian collection of microorganisms. The optimized protocol of bacilli lectins isolation and purification included precipitation with ammonium sulfate with subsequent gel filtration chromatography on Sepharose CL-6B. Hemagglutinating activity of bacilli lectins and their fine carbohydrate specificity to sialic acids and their derivatives as well as sialic asid-containing and asialic glycoconjugates were studied. The ability of extracellular bacilli lectins to discriminate a- and 13-conformation of carbohydrate molecule and the type of connection between the monomers was determined. Studied lectins showed the most affinity to glycoconjugates containing both types of sialic acids （N-acetylneuraminic acid （Neu5Ac） and N-glycolylneuraminic acid （NeuGc）） and it is supposed to be a basis of their diagnostic and analytical potential.

Application of sialic acid/polysialic acid in the drug delivery systems

The properties of modified biomaterial are gaining more and more importance in drug delivery systems.Sialic acid(SA)and polysialic acid(PSA)serve as endogenous substances,which are non-immunogenic and biodegradable.At the same time,SA modification of the drugs/carriers can enhance the uptake of tumor cell and retention in brain;PSA modifi-cation can reduce the immunogenicity of the proteins or polypeptides and increase circulation time of the modified drugs/carriers in the blood,thus achieving active targeting effect.These properties offer a variety of opportunities for applications in drug delivery systems.This article summarizes the biological functions of SA and PSA and presents the technologies of SA/PSA modified small molecule drugs,proteins and carriers in drug delivery systems.

APOPTOSIS OF EPIDIDYMIS EPITHELIUM AND CONTENT OF EPIDIDYMIS SIALIC ACID FOLLOWING TORSED/DETORSED TESTES IN RAT

Objective To investigate the apoptosis of epididyrnis epithelium and the change of epididymissialic acid following torsed/detorsed testes. Methods Twenty four adult male Sprague-Dawley rats were subjected to unilateral 720 testicular torsion with the duration of 2h and 4h, then repaired. The ischemic epididymis were collected for detecting the content of sialic acid by using spectrophotometry and the apoptosis with TUNEL technique. Results There were no statistically significant difference in the apoptosis of epididymis epithelium [(9.51± 2.78)% vs (6.34±1. 98)%] and the content of epididymis sialic acid(23, 3851 ± 9. 2199mg/mgprot vs 19. 3661 6. 3373mg/mgprot) at 24h between following 2h-torsed/detorsed testes and those of sham group. There were statistically significant difference in the apopotosis of epididymis epithelium[ (46. 81 ±3. 55)% vs (6. 34±1. 98) % ] and the content of epididymis sialic acid (13. 7249±7. 8006mg/mgprot vs 19. 3661±6. 3373mg/mgprot) at 24h between following 4h-torsed/detorsed testes and those of sham group(P <0. 05). Conclusion The results suggest that the sialic acid-secreting-function of epididymis remain normal at 24h following 2h-torsed/detorsed testes, while the apoptosis index of epididymis epithelium do not increase. The epididymis would be injured at 24h following 4h-torsed/detorsed testes, while the apoptosis index increased.

ROLE OF SIALIC ACID RESIDUES IN IRON BINDING BY HUMAN LACTOFERRIN

Lactoferrin(Lf-α)is a major constithent of the secondary granules of neutrophils, and is thought to be involved in bacteriostasis through high affinity binding of plasma Fe required for microbial growth.Lf-αis also the major Fe binding protein in milk. Fe bound to Lf-αis available for metabolic use.but the mechanism of Fe release from Lf-αis unknown.Treatment of human Lf-α with neuraminidase to remove sialic acid residues reduced its ability to bind Fe and caused release of Fe already bound to the protein. Reduction in Fe binding capacity was dependent on time of exposure to and concentration of neuraminidase,and was due to Lf-αloss or degradation. Readdition of sialic arid to desialylated Lf -a using a-2, 6-sialyltransferase restored Fe binding.The results suggest that sialylationdesialylation reactions could play a role in physiologic binding and release of Fe from the very high affinity binding sites of Lf -α.

SERUM LIPID-BOUND SIALIC ACID IN LUNG CANCER PATIENTS

Serum lipid-bound sialic acid (LSA) assay was evaluated for differential diagnosis between benign and malignant pulmonary disease.Mean serum LSA for normal controls was 13.7±3.7 mg/dl and the upper limit of the normal value was set at 19.8 mg/dl (95% confidence limit,one side).The specificity was 97.l% (95% confidence interval was 93.8%, 100.0%).The mean for benign pulmonary disease was 1 7.l±5.l mg/dl,the specificity was 89.l% (95% confidence interval was 80.9%,97.3%), and the sensitivity was 10.9%.The mean for malignant pulmonary disease was 27.l±7.9 mg/dl, and the sensitivity was 93.3%(95% confidence interval was 88.5%, 98.1%).The serum LSA levels for different malignant pulmonary diseases were different between small cell carcinoma and adenocarcinoma. The results indicate that serum LSA assay might be used as tumor marker to differentiate lung cancer from normal subjects or benign lung disease.

Correlation of Cerebrospinal Fluid Sialic Acid with Various Enzymes in Patients with Meningitis

Meningitis is often associated with cerebral compromise which may be responsible for neurological squeal in nearly half of the survivors. Little is known about the mechanisms of Central Nervous System （CNS） involvement in bacterial meningitis. The present study was undertaken to assess the changes in biochemical parameters including glucose, protein, Sialic Acid （SA） and various enzymes like Aspartate Aminotransferase （AST）, Alanine Aminotransferase （ALT）, Creatine Kinase （CK）, Gamma Glutamyltranspeptidase 7 （GGTP） and Lactate Dehydrogenase （LDH） have been determined in the Cerebrospinal Fluid （CSF） of the meningitis patients （n = 26） and compared with control subjects （n = 26）. Total Protein, CK, SA （P 〈 0.02）, AST （P 〈 0.001） and ALT （P 〈 0.0001） of meningitis patients were significantly higher, whereas, glucose （P 〈 0.05） and Gamma Glutamyltranspeptidase （GGT） （P 〈 0.01 and P 〈 0.05, respectively） were significantly lower than those of the controls. This study suggests that loss of integrity of brain-CSF barriers, sialic acid and enzyme profiles may contribute to the severity and neurological complications of meningitis.

Effect of nitrofurazone on the reproductive organs in adult male mice

Aim: To study the effect of 5-nitro-2-fumldehyde semicarbazone (nitrofurazone), a derivative of nitrofuran, onmale reproductive organs of Parkes (P) swain mice. Methods: Mice were given nitrofurazone orally at a dose of 64mg/kg body weight per day, for 10 and 20 days, and were killed 24 h and/or 56 days after the last treatment. Histo-logical appearance of testis, motility and number of spermatozoa in cauda epididymidis, and biochemical indices in epi-didymis and seminal vesicle were evaluated. Results: Histologically, testis showed marked regressive changes inthe seminiferous tubules in mice treated with nitrofurazone. Ten days after treatment, there was much depletion of germcells in the seminiferous tubules, and the germinal epithelium was lined mainly with Sertoli cells, spermatogonia, sper-matocytes, and a few round spermatids; intraepithelial vacuoles and multinucleated giant cells were also observed intubules. By 20 days, regressive changes in the seminiferous tubules were further pronounced, and pachytene spermato-cytes were the most advanced germ cells noticed in the tubules. In severe cases, the tubules were lined with a thin layerof Sertoli cells and spermatogonia. The treatment also caused marked reductions in the motility and number of sperma-tozoa in the cauda epididymidis, in weight and the level of fructose in the seminal vesicle, and in sialic acid level in theepididymis. Fifty six days after drag withdrawal, the alterations induced in the reproductive organs returned to controllevels. Conclusion: Our results suggest that nitrofurazone treatment in P mice induces marked alterations in themale reproductive organs, and that the alterations are reversible following cessation of treatment.(Asian J Androl 2001 Mar; 3: 39-44)

Clinicopathological utility of sialoglycoconjugates in diagnosing and treating colorectal cancer

Aberrant expression of glycoconjugates occurs during malignant transformation of cancer cells.Overexpression of sialoglycoconjugates in particular may play an important role in the progression,i.e.,invasion or metastasis,of cancer.Various types of sialoglycoconjugates have been investigated to clarify their biological significance and clinical utility in diagnosing and treating colorectal cancer.This review focuses specifically on expression of mucin(MUC)1 and it suggests that MUC1with the specific structure of a sialo-oligosaccharide has biological significance in determining the metastatic potential of colorectal cancer cells and clinicopathological utility in evaluating the effectiveness of treatments and the prognosis for patients with colorectal cancer.Further studies are expected to contribute to the expanded use of cancer-associated sialoglycoconjugates in cancer diagnosis and therapy.

Effect of piperine on the epididymis of adult male rats

Aim： To study the effect of piperine on the epididymal antioxidant system of adult male rats. Methods： Adult male rats were orally administered piperine at doses of 1 mg/kg, 10 mg/kg and 100 mg/kg body weight each day for 30 consecutive days. Twenty-four hours after the last treatment, the rats were weighed and killed with ether and the epididymis was dissected from the bodies. Sperm collected from the cauda region of the epididymis was used for the assessment of its count, motility and viability. Caput, corpus and cauda regions of the epididymis were separated and homogenized separately to obtain 10 % homogenates. The supernatants were used for the assays of sialic acid, superoxide dismutase, catalase, glutathione peroxidase, glutathione reductase, lipid peroxidation and hydrogen peroxide generation. Results： Body weight of the piperine-treated rats remained unchanged. The weights of the caput, corpus and cauda regions of the epididymis significantly decreased at dose of 100 mg/kg. Epididymal sperm count and motility decreased at 10 mg/kg and 100 mg/kg, and sperm viability decreased significantly at 100 mg/kg. Sialic acid levels in the epididymis decreased significantly at 100 mg/kg while significant decrease in the cauda region alone was observed at 10 mg/kg. A significant decline in the activities of superoxide dismutase, catalase, glutathione peroxidase and glutathione reductase, along with an increase in hydrogen peroxide generation and lipid peroxidation were observed at 10 mg/kg and 100 mg/kg. Conclusion： Piperine caused a decrease in the activity of antioxidant enzymes and sialic acid levels in the epididymis and thereby increased reactive oxygen species levels that could damage the epididymal environment and sperm function.

Effect of Alstonia scholaris bark extract on testicular function of Wistar rats

Abstract Aim: To evaluate the antifertility effect ofAlstonia scholaris bark extract in male rats. Methods: In male Wistar rats Alstonia scholaris bark extract was given by oral route at a dose of 200 mg/day for 60 days. The fertility and testicular function were assessed by mating tests, sperm motility, sperm concentration, biochemical indices and testicular cell population dynamics. Results: Oral feeding with the extract at a dose of 200 mg/day for the period of 60 days did not cause body weight loss, while the weights of testes, epididymides, seminal vesicle and ventral prostate were significantly reduced. The production of step-19 spermatids was reduced by 79.6% in treated rats. The population of preleptotene and pachytene spermatocytes were decreased by 61.9% and 60.1%, respectively. Spermatogonia and Sertoli cell population were also affected. The seminiferous tubule and Leydig cell nuclear area were reduced significantly (P<0.01) when compared to the controls. Reduced sperm count and motility resulted in a total suppression of fertility. A significant fall in the protein and sialic acid content of the testes, epididymides, seminal vesicle and ventral prostate as well as glycogen content of testes were also noticed. The fructose content in the seminal vesicle was lowered whereas the testicular cholesterol was elevated as compared with the controls. The following compounds were obtained from the extract with chromatographic separation over Si-gel column: α-amyrin, β-amyrin, lupiol acetate, venenative, rhazine and yohimbine. Conclusion: At the dose level employed, Alstonia scholaris bark extract has a significant antifertility effect in male rats; the primary site of action may be post meiotic germ cells (Step 19 spermatids).

Fractionation of edible bird’s nest glycoprotein hydrolysates:characterisation and antioxidative activities of the fractions

E dible bird’s nest(EBN)hydrolysates have been proven to exhibit enhanced bioactivities.However,being a macromolecule,fractions with different molecular weights would have different properties and bioactivities.Hence,this research was aimed to determine the chemical properties and antioxidant activities of freezedried(EBN_(FD))and spray-dried EBN(EBN_(SD))hydrolysates fractionated using gel permeation chromatography(GPC).Overall,two well-separated fractions were identified(EBN_(fFD1),EBN_(fFD2),EBN_(fSD1) and EBN_(fSD2)).EBNFD demonstrated significantly higher(P≤0.05)peptide(3.6%),total carbohydrate(27.7%)and sialic acid(18.2%)contents than that of EBNSD.Similar trend was observed in low molecular weight fractionates(EBN_(fFD2) and EBN_(fSD2)).Meanwhile,the first fractionates(EBN_(fFD1) and EBN_(fSD1))exhibited significantly higher(P≤0.05)hydroxyl radical(·OH)scavenging activity.Fourier transform infrared(FTIR)spectroscopy demonstrated that all EBN fractionates have similar spectrum,except in the region of N—H(amideⅡ)and C—H alkyl group.In conclusion,EBN fractionates with different molecular weights showed different chemical properties and antioxidant activities.

N-Propionylmannosamine stimulates axonal elongation in a murine model of sciatic nerve injury

Increasing evidence indicates that sialic acid plays an important role during nerve regeneration. Sialic acids can be modified in vitro as well as in vivo using metabolic oligosaccharide engineering of the N-awl side chain. N-Propionylmannosamine （ManNProp） increases neurite outgrowth and accelerates the reestablishment of functional synapses in vitro. We investigated the influence of systemic ManNProp application using a specific in vivo mouse model. Using mice expressing axonal fluorescent proteins, we quantified the extension of regenerating axons, the number of regenerating axons, the number of arborising axons and the number of branches per axon 5 days after injury. Sciatic nerves from non-expressing mice were grafted into those expressing yellow fluorescent protein. We began a twice-daily intraperitoneal application of either peracetylated ManNProp （200 mg/kg） or saline solution 5 days before injury, and continued it until nerve harvest （5 days after transection）. ManNProp significantly increased the mean distance of axonal regeneration （2.49 mm vs. 1.53 mm; P 〈 0.005） and the number of arborizing axons （21% vs. 16%; P = 0.008） 5 days after sciatic nerve grafting. ManNProp did not affect the number of regenerating axons or the number of branches per arborizing axon. The biochemical glycoengineering of the N-acyl side chain of sialic acid might be a promising approach for improving peripheral nerve regeneration.

Effect of papaya seed extract on microenvironment of cauda epididymis

Aim: To evaluate the effect of aqueous Carica papaya seed extract on microenvironment of cauda epididymis.Methods: Adult male albino rats were intramuscularly administered with 0 (control) or 0.5 mg papaya seed ex-tract/kg body weight for 7 days. Cauda epididymal tubular content was collected by micropuncture technique; epididy-mal luminal fluid and sperm pellets were separately analyzed. Results: The results revealed that the extract treat-ment caused significant reduction, as compared with control, in total protein and sialic acid contents in both epididymalfluid and sperm pellet. As compared with control, significantly lowered acid phosphatase activity was recorded in spermpellet but was higher in epididymal fluid after the treatment. The extract treatment also caused significant reduction inlevel of inorganic phosphorus in the epididymal fluid. Conclusion: It is concluded that the aqueous papaya seed ex-tract alters cauda epididymal microenvironment.

Inhibition of Probimane on Lipid Peroxidation of Rabbit and Human Erythrocytes

Lipid peroxide (LPO) plays pivotal roles in the process and development of many diseases. In this work, we studied the inhibitory effect of probimane (Pro), a Chinese anticancer agent, on erythrocyte LPO and the interaction of Pro with sialic acids (sia). Malondialdehyde (MDA) of erythrocytes activated by hydrogen peroxide was measured. Pro was found to inhibit the product of LPO induced by hydrogen peroxide in a non-enzyme system of both rabbit and human erythrocytes in the absence of doxorubicin. Sia were found to enhance LPO production and the activity of N-glycolylneuraminic acid (NeuGc) was about 5 times higher than that of N-acetylneuraminic acid (5AcNeu) at equivalent concentrations. Pro inhibited the increased LPO production induced by sia and the activity of Pro against LPO with 5AcNeu was almost twofold higher than that of Pro alone. It suggests that Pro be an inhibitor of LPO (free radicals) and as a functional modulator of sia in body.

Sialic acid-mediated photochemotherapy enhances infiltration of CD8^(+)T cells from tumor-draining lymph nodes into tumors of immunosenescent mice

Immunoscenescence plays a key role in the initiation and development of tumors.Furthermore,immunoscenescence also impacts drug delivery and cancer therapeutic efficacy.To reduce the impact of immunosenescence on anti-tumor therapy,this experimental plan aimed to use neutrophils with tumor tropism properties to deliver sialic acid(SA)-modified liposomes into the tumor,kill tumor cells via SA-mediated photochemotherapy,enhance infiltration of neutrophils into the tumor,induce immunogenic death of tumor cells with chemotherapy,enhance infiltration of CD8^(+)T cells into the tumordraining lymph nodes and tumors of immunosenescent mice,and achieve SA-mediated photochemotherapy.We found that CD8^(+)T cell and neutrophil levels in 16-month-old mice were significantly lower than those in 2-and 8-month-old mice;16-month-old mice exhibited immunosenescence.The anti-tumor efficacy of SA-mediated non-photochemotherapy declined in 16-month-old mice,and tumors recurred after scabbing.SA-mediated photochemotherapy enhanced tumor infiltration by CD8^(+)T cells and neutrophils,induced crusting and regression of tumors in 8-month-old mice,inhibited metastasis and recurrence of tumors and eliminated the immunosenescence-induced decline in antitumor therapeutic efficacy in 16-month-old mice via the light-heat-chemical-immunity conversion.

Inhibition of Probimane on Lipoperoxidation of Human Red Cells in Vitro

To investigate the influence of probimane on the lipoperoxidation of human red cells in vitro. Malondialdehyde (MDA) of human red cells induced by hydrogen peroxide was assayed by means of colorimetry. It was shown that probimane ( > 2 μM ) could inhibit the lipoperoxidation without adriamycin. The inhibitory rates of probimane were 9.3 % ( 2 μM ) and 35.3 % ( 10 μM ) respectively ( P< 0.05). Probimane ( 2 μM ) inhibited lipoperoxidation in the presence of sialic acid ( 0.4 μM ) ( inhibitory rate 17.8 % ) was approximately two fold higher than those without sialic acid ( inhibitory rate 9.3 % ). So probimane may inhibit tumor growth through affecting lipoperoxidations and exhibit anticancer effects via sialic acids.

THE ENHANCEMENT OF TWEEN-80 ON THE ANTITUMOR EFFECT OF THE HYPERTHERMIA41℃ IN TUMOR-BEARING MICE

B16 melanoma cells were inoculated into the BALB/C mice to establish melanoma-bearing models. The antitumor effect of Tween-80 in combination with hyperthermia 41℃ was studied. We observed the changes of the mortality of tumor-bearing mice, the tumor growth curves, activities of serum tumor necrosis factor (STNF) and the level of serum sialic acid (SSA) in tumor-bearing mice. The number of pulmonary metastatic tumor foci from blood flow was also detected. The results showed that combined with Twecn-80, hyperthermia 41℃ could dramatically suppress the growth of the melanoma in the feet of mice, survive the tumor-bearing mice and decrease the number of pulmonary metastatic tumor foci but no significant effects were observed by treatment with Tween-80 or hyperthermia at 41℃ alone. The activities of STNF and the level of SSA of the melanoma-bearing mice kept at higher levels than those of normal BALB/C mice. Tween-80 combined with heating 41℃ significantly decreased activities of STNF, but the level of SSA still kept high level within 1 to 2 weeks and decreased 10 weeks later with the tumor regression. These results demonstrate that Tween-80 may make hyperthermia exert effective antitumor effect below the critical temperature and increase the safety of hyperthermia in treatment. It is one of the most ideal synergist with hyperthermia. The changes of STNF and SSA suggest that the synergeticeffect could involve in facilitating the exposure of tumor antigen and activation of immune system.

Biochemical composition of the glomerular extracellular matrix in patients with diabetic kidney disease

In the glomeruli,mesangial cells produce mesangial matrix while podocytes wrap glomerular capillaries with cellular extensions named foot processes and tether the glomerular basement membrane(GBM).The turnover of the mature GBM and the ability of adult podocytes to repair injured GBM are unclear.The actin cytoskeleton is a major cytoplasmic component of podocyte foot processes and links the cell to the GBM.Predominant components of the normal glomerular extracellular matrix(ECM)include glycosaminoglycans,proteoglycans,laminins,fibronectin-1,and several types of collagen.In patients with diabetes,multiorgan composition of extracellular tissues is anomalous,including the kidney,so that the constitution and arrangement of glomerular ECM is profoundly altered.In patients with diabetic kidney disease(DKD),the global quantity of glomerular ECM is increased.The level of sulfated proteoglycans is reduced while hyaluronic acid is augmented,compared to control subjects.The concentration of mesangial fibronectin-1 varies depending on the stage of DKD.Mesangial type III collagen is abundant in patients with DKD,unlike normal kidneys.The amount of type V and type VI collagens is higher in DKD and increases with the progression of the disease.The GBM contains lower amount of type IV collagen in DKD compared to normal tissue.Further,genetic variants in theα3 chain of type IV collagen may modulate susceptibility to DKD and end-stage kidney disease.Human cellular models of glomerular cells,analyses of human glomerular proteome,and improved microscopy procedures have been developed to investigate the molecular composition and organization of the human glomerular ECM.