Serum Levels and Glycosylation Changes of Alpha-1-Acid Glycoprotein According to Severity of Breast Cancer in Korean Women.

Elevated serum levels of alpha-1-acid glycoprotein (AGP) are known to be associated with several types of cancer. In addition, some reports have indicated that changes in glycosylation of AGP are associated with cancer progression. However, changes in AGP levels of serum and changes in glycosylation of AGPs in breast cancer have not been specifically studied. In the present study, serum AGP levels in benign (BN) cancer and breast cancer stage I (BC I), BC IIA, BC IIB, and BC III in Korean women were measured using an enzyme-linked immunosorbent assay (ELISA). AGP was purified from individual sera by hot phenol extraction and then subjected to AGP glycosylation analysis. Three types of AGP glycosylation (fucosylation, high-mannose-type and sialylation) were detected using enzyme-linked lectin assays (ELLAs). Serum AGP levels were higher in BC I, BC IIA, BC IIB, and BC III, than in the BN group, and the level in BC I and BC IIA was high enough to be distinguished from BN. Meanwhile, terminal fucosylation and high-mannose-type glycans appeared to be lowest in BC I. The glycosylation levels of BC I provide sensitivity and specificity that make BC I clearly distinguishable from BC IIA, BC IIB, and BC III as well as BN. Therefore, determination of serum AGP or AGP glycosylation level could be useful for detecting the early stages of breast cancer.

Current status and future directions of fish vaccines employing virus-like particles.

In most breeding schemes, fish are cultured in enclosed spaces, which greatly increases the risk of outbreaks where the onset of infectious diseases can cause massive mortality and enormous economic losses. Vaccination is the most effective and long-term measure for improving the basic make-up of a fish farm. As the relationship between antibody and antigen is similar to that between screw and nut, similarity in the shape or nature of the vaccine antigen to the original pathogen is important for achieving a satisfactory/good/excellent antibody response with a vaccine. Virus-like particles (VLPs) best fulfil this requirement as their tertiary structure mimics that of the native virus. For this reason, VLPs have been attracting attention as next-generation vaccines for humans and animals, and the effects of various types of VLP vaccines on humans and livestock have been examined. Recent studies of VLP-based fish vaccines indicate that these vaccines are promising, and raise hopes of extending their use in the near future. In this review, the structural properties and immunogenicity of VLP-based vaccines against fish viruses such as infectious pancreatic necrosis virus (IPNV), salmonid alphavirus (SAV), nervous necrosis virus (NNV) and iridovirus are introduced/summarized. The NNV VLP vaccine is the most-studied VLP-based vaccine against fish viruses. Therefore, the current status of NNV VLP research is highlighted in this review, which deals with the advantages of using VLPs as vaccines, and the expression systems for producing them. Moreover, the need for lyophilized VLPs and oral VLP delivery is discussed. Finally, future directions for the development of VLP vaccines in the fish vaccine field are considered.

IgG and IgM responses to human papillomavirus L1 virus-like particle as a function of dosing schedule and vaccine formulation.

Most commercialized virus-like particle (VLP) vaccines use aluminum salt as adjuvant, even though VLPs provoke adequate antibody responses without adjuvant. We do not have detailed knowledge of how adjuvant affects the profile of anti-VLP antibodies. Meanwhile, there is evidence that differences between vaccination protocols influence the glycosylation of antibodies, which may alter their effector functions. In the present study a murine model was used to investigate the effects of dosing schedule and adjuvant on the antibody profiles and glycosylation levels of antigen-specific antibody responses to human papillomavirus type 16 L1 (HPV16 L1) VLPs. Mice received subcutaneously 2,000 ng of antigen divided into 4 or 7 doses. The HPV16 L1 VLPs elicited > 4 log10 anti-HPV16 L1 IgG titers without adjuvant, and aluminum hydroxide as adjuvant increased IgG titers 1.3- to 4-fold and reduced the anti-HPV16 L1 IgG2a / anti-HPV16 L1 IgG1 ratio value (use of aluminum hydroxide reduced the ratio of the IgG2a). Immunization with HPV16 L1 VLPs in combination with Freund's adjuvant enhanced IgG titers 5- to 12-fold. Seven-dose immunization markedly increased anti-HPV16 L1 IgM titers compared to four-dose immunization, as well as increasing the proportion of glycosylated antibodies. Our results suggest that antibody glycosylation can be controlled immunologically, and IgG and IgM profiles and glycosylation profiles of the vaccine-induced antibodies can be used as indicators reflecting the vaccine characteristics. These results indicate that the HPV16 L1 VLP dosing schedule can affect the quality of antigen-specific antibody responses. We suggest that dosing schedules should be noted in vaccination protocols for VLP-based vaccines.

Serum anti-GAPDH autoantibody levels reflect the severity of cervical lesions: A potential serum biomarker for cervical cancer screening.

Recent studies have indicated that a certain level of autoantibodies may be essential for maintaining good health as well as preventing cancer development, and that the levels of serum autoantibodies can decline during malignant progression. The aim of the present study was to identify such an autoantibody-based biomarker for screening cervical lesions. An autoantigen reactive with healthy female sera was detected in the cytosolic fraction of HeLa cells, a cervical cancer cell line, and identified. Serum immunoglobulin (Ig)-G and IgM levels against the purified autoantigen in normal, cervical intraepithelial neoplasias (CINs) I, II and III, and cervical cancer were compared using ELISAs. The autoantigen in HeLa cells was identified to be GAPDH. The serum levels of anti-HeLa-GAPDH IgG decreased with increasing severity of cervical lesions, and similar decreases in IgM levels were revealed. Notably, the anti-HeLa-GAPDH IgG level was discovered to discriminate cervical cancer from normal samples with 80.0% sensitivity and 96.6% specificity. The serum anti-HeLa-GAPDH autoantibody level, as a single parameter, is a promising serum biomarker for screening cervical lesions.

Profiling of serum antibodies against human papillomavirus antigens in Korean women with cervical intraepithelial neoplasia and cervical cancer.

Sero-epidemiological studies of human papillomavirus (HPV) have been undertaken over the last two decades. In this study, the prevalences of nine serum antibodies (anti-E6, E7 and L1 antibodies of HPV types 16, 18, and 58) were evaluated in normal (control) Korean women and women with cervical intraepithelial neoplasia (CIN) I, CIN II, CIN III, and cervical cancer. The frequencies of all types of anti-HPV antibodies were higher in the CIN stages and cervical cancer than in normal women, and those of anti-HPV16 E6 and E7, anti-HPV18 E6 and E7, and anti-HPV58 E7 antibodies were higher in the cervical cancer group than in the CIN stages. The frequencies of antibodies against HPV16, 18, and 58 E7 tended to increase with increasing severity of cervical lesions. However, there were few differences in the frequencies of antibodies against the L1 antigens of HPV16, 18 and 58 in cervical cancer versus CIN stages. The anti-HPV antibodies were detected in 26.5% of normal, 46.3% of CIN I, 62.5% of CIN II, 51.6% of CIN III, and 75% of cancers when any of the nine antigens was used as a criterion. Correlations between HPV DNA positivity and seropositivity for anti-HPV E6, E7, or L1 antibodies were found only in HPV16 DNA-positive cervical cancers for anti-HPV16 E6 and L1 antibodies. In addition, strong positive correlations in seropositivity were found between anti-HPV16 E7 and anti-HPV58 E7 antibodies, and between anti-HPV18 E6 and anti-HPV58 E6 antibodies. These findings should advance global profiling of the seroprevalences of antibodies against HPV antigens.

Use of CA15­3 for screening breast cancer: An antibody­lectin sandwich assay for detecting glycosylation of CA15­3 in sera.

Elevated serum CA15­3 assessed by enzyme­linked immunosorbent assay (ELISA) has been considered a diagnostic marker of breast cancer. However, accumulating data indicate that the current ELISA system for detecting CA15­3, which targets the peptide backbone of CA15­3, is not sufficiently sensitive to detect early or localized breast cancer. In the present study, we designed an antibody­lectin sandwich assay detecting glycosylation of CA15­3 in patients with breast cancer. Ιmmobilized anti­CA15­3 monoclonal antibody captures CA15­3 in serum, and glycosylation of the CA15­3 is detected with Concanavalin A (ConA) lectin, which preferentially bind high­mannose N­glycans. ConA provided the best signal for detecting serum CA15­3 among 9 types of lectin, Since CA15­3 is a heavily glycosylated protein, detecting the glycosylation of CA15­3 should be a much more sensitive way to assess CA15­3 than the current ELISA method. Linear responses were obtained in the anti­CA15­3 antibody­ConA sandwich assay when sera were diluted up to 2000­fold. This dilution factor is comparable with that of the current ELISA system which allows 50­ to 100­fold serum dilutions. The glycosylation level of CA15­3 was found to increase with increasing breast cancer stage in the sandwich assay. The assay system appeared to efficiently discriminate breast cancer stage I (sensitivity: 63%, specificity: 69%), IIA (sensitivity: 77%, specificity: 75%), IIB (sensitivity: 69%, specificity: 86%) and III (sensitivity: 80%, specificity: 65%) from benign breast disease. The antibody­lectin sandwich assay shows promise as a new prospect for the early detection of breast cancer.

Comparison of the size distributions and immunogenicity of human papillomavirus type 16 L1 virus-like particles produced in insect and yeast cells.

Insect and yeast cells are considered the expression systems of choice for producing virus-like particles (VLPs), and numerous types of VLPs have been produced in these systems. However, previous studies were restricted to identifying the characteristics of individual VLP preparations. No direct comparison of the structures and immunogenic properties of insect and yeast-derived VLPs has so far been made. In the present study, the size distribution and immunogenic properties of human papillomavirus type 16 (HPV16) L1 VLPs produced in Spodoptera frugipedra-9 insect cells and Saccharomyces cerevisiae were compared. The insect cell-derived VLPs were larger than the yeast ones (P < 0.0001), with median sizes of 34 and 26 nm, respectively. In addition, the insect-derived VLPs appeared to be more diverse in size than the yeast-derived VLPs. Immunization of mice with 30 ng per dose of VLPs elicited 2.7- and 2.4-fold higher anti-HPV16 L1 IgG and anti-HPV16 neutralizing antibody titers than immunization with the same amounts of the yeast-derived VLPs after the 4th immunizations, respectively. Our results suggest that the choice of expression system critically affects the particle size and immunogenic property of HPV16 L1 VLPs.

Use of protein-based biomarkers of exfoliated cervical cells for primary screening of cervical cancer.

There has been no attempt to apply protein-based markers of exfoliated cervical cells (ECCs) for primary screening of cervical cancer. In the present study, the levels of six tumor-associated protein [TAPs: Sialyl Lewis A (SLeA), Cancer Antigen 15-3 (CA 15-3), p53, heat shock protein (Hsp)70, Hsp27 and squamous cervical carcinoma antigen (SCCA)]and of two human papillomavirus (HPV) viral proteins (HPV16 E7 and HPV16 L1) of ECCs lysates were evaluated by enzyme-linked immunosorbent assays (ELISAs).The wells of 96-well plates were coated with the ECCs lysates from normal, cervical intraepithelial neoplasia (CIN) I, CIN II, CIN III and cancer groups, and candidate proteins were detected by relevant antibodies. SLeA level decreased with increasing severity of lesions, whereas the levels of other candidate proteins increased. SLeA, HPV16 L1 and p53 levels appeared more useful for detecting cervical lesions than the other candidates. The combination of ELISA-SLeA and ELISA-HPV16 L1 could efficiently detect cervical lesions from normal. The combination of ELISA-SLeA and ELISA-p53 could powerfully discriminate cancer from normal with 91.3% sensitivity and 96.7% specificity. The protein levels of ECCs have great potential as biomarkers for primary screening of cervical cancer.

Stability of virus-like particles of red-spotted grouper nervous necrosis virus in the aqueous state, and the vaccine potential of lyophilized particles.

Virus-like particles (VLPs) are multi protein complexes mimicking the structural properties of the native virus. The development of freeze-dried formulations of such complex protein structures remains a challenge. Red-spotted grouper nervous necrosis virus (RGNNV) causes mass mortality in fish culture, and RGNNV VLPs have been suggested to be promising vaccine candidates. In the present study, the stability of RGNNV VLPs in the liquid state was investigated over a 4-week period, along with the influence of freeze-drying on VLP stability. RGNNV VLPs were completely degraded after one week at 37 °C followed by 3 weeks at ambient temperature, and they were partially degraded after 4 weeks at 4 °C. Therefore, the inherent stability of RGNNV VLP in an aqueous milieu is insufficient for long-term storage. When RGNNV VLPs were freeze-dried in the presence or absence of sugar stabilizers, sorbitol was found to improve VLP stability whereas mannitol reduced it. VLP preparations freeze-dried with sorbitol or without stabilizer were as immunogenic as control (non-freeze dried) VLPs, whereas VLPs freeze-dried in mannitol were less immunogenic. These results indicate that freeze-dried RGNNV VLPs have potential as vaccines.

Use of autoantibodies against tumor-associated antigens as serum biomarkers for primary screening of cervical cancer.

Serum autoantibodies against tumor-associated antigens (TAAs) have received much attention as potential biomarkers for early detection of cancers, since they can be detected in the early stages of cancers. Autoantibodies against Cancer Antigen 15-3 (CA15-3), carcinoembryonic antigen (CEA), Cancer Antigen 19-9 (CA19-9), c-Myc, p53, heat shock protein (Hsp)27 and Hsp70 have been suggested as potential markers for detecting several types of cancer. In the present study, the seven types of antibody listed above were evaluated for detecting cervical lesions. Enzyme-linked immunosorbent assays (ELISAs) were used to measure IgG levels of the autoantibodies in women with normal cytology, cervical intraepithelial neoplasia (CIN) I, CIN II, CIN III and cervical cancer. The increases of anti-CA15-3 and anti-CEA IgG in cervical cancer were more pronounced than the increases of the other markers, and the level of anti-CA19-9 IgG in CIN III stage was higher than in normal CIN I, CIN II or cervical cancer. A combination of ELISAs detecting anti-CA15-3, anti-CEA and anti-CA19-9 IgGs was found to reliably discriminate CINs from normal and to strongly differentiate cancer from normal (90.3% of sensitivity and 82.1% of specificity). We suggest that the combination of three ELISA may be useful for detecting cervical lesions.

Current status and future prospects for human papillomavirus vaccines.

Cervical cancer is the fourth most frequent cancer among women worldwide. Human papillomaviruses (HPVs) cause almost all cervical cancers in low-income countries. Three prophylactic HPV virus-like particle-based vaccines have been licensed to date, and they have all shown high efficacy and reliable safety profiles. However, isolated safety issues have resulted in a reluctance to use these vaccinations. In addition, the high prices of the vaccinations have caused the inequitable distribution of the vaccine: the prices are unaffordable for low-income countries. Meanwhile, great effort has been put into the development of therapeutic HPV vaccines, including protein/peptide-, live vector-, DNA- and cell-based vaccines. These new vaccines have considerable therapeutic potential but limited practical use. The development of immune checkpoint inhibitors and personalized immunotherapy remain challenges for future study. In this article, the current status of the licensed vaccines, therapeutic HPV vaccines and biosimilars, and new platforms for HPV vaccines, are reviewed, and safety issues related to the licensed vaccines are discussed. In addition, the prospects for HPV vaccines are considered.

Oral vaccination through voluntary consumption of the convict grouper Epinephelus septemfasciatus with yeast producing the capsid protein of red-spotted grouper nervous necrosis virus.

Nervous necrosis viruses (NNV) cause serious economic losses in marine fish cultivation. The red-spotted grouper NNV (RGNNV) is the most common species of NNV worldwide. There have been many efforts to develop prophylactic NNV vaccines, and various types of vaccine candidate have been suggested. However, most were designed as injectable vaccines, which are not suitable for large-scale vaccination and cause too much stress to the fish. Oral vaccination through voluntary feeding is an ideal way to provide protective immunity to fish. In the present study, recombinant Saccharomyces cerevisiae producing RGNNV capsid protein was used as oral vaccine. The recombinant yeast was prepared in freeze-dried form after disruption. Convict groupers were divided into three groups, control, and oral and parenteral vaccination groups, each consisting of 700 fishes. The control group received no treatment, the parenteral group received one intraperitoneal injection of RGNNV virus-like particles, and the oral vaccination group consumed feed containing the lysed recombinant yeast; voluntary intake was allowed four times at one-week intervals. Both vaccination groups produced serum RGNNV neutralizing antibody titers of >103 (log 2, 9.96), sustained for at least 95days post-immunization. In addition, in response to challenge with RGNNV both groups suffered significantly reduced mortality and had reduced brain RGNNV titers. These results indicate that recombinant yeast-based oral fish vaccines have great potential for large-scale vaccination.

Two-step chromatographic purification of glutathione S-transferase-tagged human papillomavirus type 16 E6 protein and its application for serology.

Human papillomavirus (HPV) E6 protein is an oncoprotein with a pivotal role in cervical carcinogenesis. Expression and purification of HPV E6 from Escherichia coli (E. coli) has been difficult because of its strong hydrophobicity even when expressed as a fusion protein with glutathione S-transferase (GST). There has been no protocol suggested for purifying GST-tagged HPV E6 protein with high purity so far. Herein, we provide efficient protocol for purifying GST-HPV16 E6 protein for the first time. In the current study, the GST-tagged protein was expressed in E. coli and a purification method was designed using cation-exchange chromatography followed by GST-affinity chromatography. Using physiological pH buffer during cell lysis and first cation-exchange chromatography significantly reduced yield of full-length GST-HPV16 E6 protein. It was found that using an alkaline buffer during cation-exchange chromatography was needed to obtain full length GST-HPV16 E6 protein. GST-HPV16 E6 protein recovered from the purification using alkaline condition retained its inherent p53-binding ability. Moreover, we were able to detect anti-HPV16 E6 antibodies with high sensitivity in sera from patients with cervical cancer using the GST-HPV16 E6 protein. It was found that the GST-HPV16 E6 protein could be used as a coating agent to enhance the sensitivity of detection of serum anti-HPV16 E6 antibodies when treated with ethylene glycol-bis (ß-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA). These results indicate that the two-step chromatographic purification allows obtaining high purity of GST-HPV16 E6 protein and the GST-HPV16 E6 is suitable to be used as an antigen of serology assay.

Characterization of human papillomavirus type 16 pseudovirus containing histones.

BACKGROUND: Pseudoviruses (PsVs) that encapsidate a reporter plasmid DNA have been used as surrogates for native human papillomavirus (HPV), whose continuous production is technically difficult. HPV PsVs have been designed to form capsids made up of the major capsid protein L1 and the minor capsid proteins L2. HPV PsVs have been produced in 293TT cells transfected with plasmid expressing L1 and L2 protein and plasmid containing the reporter gene. Several studies have suggested that naturally occurring HPV virions contain cellular histones, and histones have also been identified in mature HPV PsVs. However, the effect of the histones on the properties of the PsVs has not been investigated. Using heparin chromatography, we separated mature HPV type 16 PsVs into three fractions (I, II, and III) according to their heparin-binding affinities. RESULTS: The amounts of cellular histone and cellular nucleotides per PsV were found to increase in the order fraction I, II and III. It appeared that PsVs in fraction I contains just small amount of cellular histone in Western blot analysis. The proportions of the three fractions in PsV preparations were 83.4, 7.5, and 9.1 % for fraction I, II, and III PsVs, respectively. In the electron microscope PsVs in fraction I appeared to have a more condensed structure than those in fractions II and III. Under the electron microscope fraction II and III PsVs appeared to be covered by substantial amounts of cellular histone while there was no visible histone covering PsVs of fraction I. Also the levels of reporter gene expression in infections of fraction II and III PsVs to 293TT cells were significantly lower than those in infections of fraction I PsV, and fraction II and III particles had significantly reduced immunogenicity. CONCLUSIONS: Our findings suggest that the involvement of large amounts of cellular histones during PsV formation interferes with the structural integrity of the PsVs and affects their immunogenicity. The fraction I particle therefore has the most suitable characteristics for use as an HPV PsV.

Effects of carboxypeptidase B treatment and elevated temperature on recombinant monoclonal antibody charge variants in cation-exchange chromatography analysis.

Charge variants (acidic and basic) of recombinant monoclonal antibodies (Mabs) have received much attention due to their potential biological effects. C-terminal lysine variants are common in Mabs and their proportion is affected by the manufacturing process. In the present study, changes of trastuzumab charge variants brought about by carboxypeptidase B treatment and subsequent storage at 8 or 37 °C for up to 24 h were monitored by cation-exchange chromatography analysis to investigate the effects of C-terminal lysine cleavage and its subsequent reaction at 8 or 37 °C. C-terminal lysine cleavage at 8 °C reduced the fraction of basic species and had little effect on the fraction of acidic species. Analysis of individual peaks demonstrated that C-terminal lysine cleavage induced both increases and decreases in individual acidic variants, with the result that there was little overall change in the overall proportion of acidic species. It appeared that most of the basic variant Mab molecules but only a fraction of the acidic variant molecules had C-terminal lysines. Increasing the temperature to 37 °C appeared to increase the fraction of acidic species and decrease main species significantly, without a similar change in basic species. These results indicate that length of exposure to elevated temperature is a critical consideration in charge variant analysis.

A lectin-based approach to detecting carcinogenesis in breast tissue.

It has been suggested that the diversity of glycosylation structures that form during cancer progression and the sensitivity with which they are able to be detected have great potential for cancer screening. However, the large majority of breast cancer research has instead focused on the development of protein or nucleic acid markers. In the present study, alterations in glycosylation in breast cancer tissue were analyzed using enzyme-linked lectin assays (ELLAs), which have potential for high-throughput screening. Cancer tissues (CCs) and normal tissues (CNs) were collected from women with breast cancer ranging from stage 0 to IIIA. The specimens were divided into two groups, stage 0-I and stage II-III, and the levels of four types of lectin in stage 0-I and stage II-III CCs and CNs were compared by ELLA. The results demonstrated that, relative to CNs, the CCs contained significantly enhanced levels of mannosylation (stage 0-I, P<0.001; stage II-III, P<0.001), galactosylation (stage 0-I, P<0.05; stage II-III, P<0.001), sialylation (stage 0-I, P<0.001; stage II-III, P<0.01) and fucosylation (stage 0-I, P<0.01; stage II-III, P<0.01). Furthermore, stage II-III CCs had higher levels of mannosylation (P<0.05) and galactosylation (P<0.01) than stage 0-I CCs. The sensitivity of the ELLA system ranged from 71-100% when specificity was set at 100%. These results demonstrate that enhanced glycosylation levels identified by ELLA are associated with the development of breast tumors, and provide evidence of the exceptional sensitivity and specificity of the ELLA system in the detection of breast cancer. This approach is anticipated to contribute highly to the development of reliable diagnostic procedures for breast cancer.

Increased sialylation and reduced fucosylation of exfoliated cervical cells are potential markers of carcinogenesis in the cervix.

BACKGROUND: The Pap test has been used for over 50 years for primary screening of cervical cancer. There has been no study of glycosylation changes in Pap test samples despite considerable potential of the glycosylation changes as biomarkers for detecting cancerous lesions. In this study, we developed a 96-well platform for enzyme-linked lectin assays (ELLAs) to evaluate glycosylation levels in cervical cells. METHODS: A total of 117 samples of exfoliated cervical cells (ECCs) from 37 individuals with normal cytology, 20 with cervical intraepithelial neoplasia (CIN) 1, 19 with CIN 2, 26 with CIN 3 and 15 with cervical cancer were analyzed by ELLAs. The wells of 96-well plates were coated with lysates of the cervical cells, and sialylation and fucosylation levels were compared between the groups. RESULTS: Sialylation levels increased and fucosylation levels decreased with increasing grade of cervical dysplasia. ELLAs for sialylation [ELLA-Sambucus nigra (SNAs)] and fucosylation [ELLA-Aleuria aurantia lectin (AAL)] discriminated not only CIN 2 and worse (CIN 2+: CIN 2, CIN 3, and cancer) from normal cytology but also CIN 3 and worse (CIN 3+: CIN3 and cancer) from normal cytology. ELLA-SNAs and ELLA-AALs distinguished cancer from normal cytology with a high true-positive rate (TPR) (ELLA-SNAs: 87%; ELLA-AALs: 87%) and low false-positive rate (FPR) (ELLA-SNAs: 19%; ELLA-AALs: 11%). CONCLUSIONS: The sialylation and fucosylation levels of ECCs as measured by ELLAs have great potential as biomarkers for primary screening of cervical cancer.

Ameliorating Effect of Dietary Xylitol on Human Respiratory Syncytial Virus (hRSV) Infection.

Human respiratory syncytial virus (hRSV) is the most common cause of bronchiolitis and pneumonia in infants. The lack of proper prophylactics and therapeutics for controlling hRSV infection has been of great concern worldwide. Xylitol is a well-known sugar substitute and its effect against bacteria in the oral cavity is well known. However, little is known of its effect on viral infections. In this study, the effect of dietary xylitol on hRSV infection was investigated in a mouse model for the first time. Mice received xylitol for 14 d prior to virus challenge and for a further 3 d post challenge. Significantly larger reductions in lung virus titers were observed in the mice receiving xylitol than in the controls receiving phosphate-buffered saline (PBS). In addition, fewer CD3(+) and CD3(+)CD8(+) lymphocytes, whose numbers reflect inflammatory status, were recruited in the mice receiving xylitol. These results indicate that dietary xylitol can ameliorate hRSV infections and reduce inflammation-associated immune responses to hRSV infection.

Age-related reduction of antibody response against the human endogenous retrovirus K envelope in women.

In the present study, the correlation between the antibody response against human endogenous retrovirus K (HERV-K) envelope and human age was investigated. Antibody levels were compared in groups in their 20s (n = 25), 30s (n = 39), 40s (n = 68), 50s (n = 32), and 60s and over (n = 25), which included healthy individuals and breast cancer and/or cervical cancer patients. It appeared that both IgM and IgG responses against the HERV-K envelope fell with increasing age. There were no differences in anti-HERV-K envelope antibody levels between healthy individuals and cancer patients. Therefore, our results indicated that the anti-HERV-K antibody levels cannot be considered as cancer-specific marker. Also, IgG1 appeared to be the predominant subtype in the reduction of the IgG response by age. Receiver operating characteristic curves of anti-HERV-K envelope IgM levels indicated that the groups of people in their 20s or 30s could be distinguished from those in their 40s, 50s or 60s and over with satisfactory sensitivity and specificity. These findings indicate that the serum antibody level of HERV-K envelope is a critical parameter reflecting person's age.

A lectin-based diagnostic system using circulating antibodies to detect cervical intraepithelial neoplasia and cervical cancer.

In the present study, we developed serological strategies using immunoglobulin fractions obtained by protein A chromatography to screen for cervical cancer and cervical intraepithelial neoplasia I (CIN I). The reactivities of the immunoglobulins purified from sera of women with normal cytology, CIN I and cervical cancer were compared in enzyme-linked immunosorbent assays (ELISA) and enzyme-linked lectin assays (ELLAs). To capture the immunoglobulins, ELISAs and ELLAs were performed in protein A immobilized microplates. The reactivity of immunoglobulin in ELISA was in the increasing order normal cytology, CIN I and cervical cancer, while that in ELLAs for detecting fucosylation was in the decreasing order normal cytology, CIN I and cervical cancer. It was confirmed that women with CIN I were distinguishable from women with normal cytology or women with cervical cancer in the ELISA or the ELLA for detecting fucosylation with considerable sensitivity and specificity. Women with cervical cancer were also distinguishable from women with normal cytology with high sensitivity (ELISA: 97%, ELLA: 87%) and specificity (ELISA: 69%, ELLA: 72%). Moreover, the logistic regression model of the ELISA and the ELLA discriminated cervical cancer from normal cytology with 93% sensitivity and 93% specificity. These results indicate that the ELISAs and the ELLAs have great potential as strategies for primary screening of cervical cancer and CIN. It is expected that the ELISA and the ELLA can provide new insights to understand systemic changes of serum immunoglobulins during cervical cancer progression.