Total Serum Glycoprotein Research Articles

Impact of estrogen on IgG glycosylation and serum protein glycosylation in a murine model of healthy postmenopause.

The glycosylation of immunoglobulin (Ig) G regulates IgG interaction capability with Fc gamma receptors found in all immune cells. In pathogenic conditions, estrogen can impact IgG levels and glycosylation. Following menopause, when estrogen levels decline affecting the immune system and potentially leading to a heightened susceptibility of immune activation. In this study, we aim to determine if estrogen levels can regulate IgG glycosylation in postmenopausal healthy situations. Mice were ovariectomized to simulate an estrogen-deficient postmenopausal status and then treated with 17-beta-estradiol (E2) at different doses and different administration strategies. Using a highly sensitive liquid chromatography-tandem mass spectrometry (MS/MS) glycoproteomic method, we demonstrated that E2 treatment increased the degree of glycosylation on IgG-Fc with both galactosylation and sialylation in the position required for interaction with Fc gamma receptors. We also observed that only long-term estrogen deficiency reduces IgG levels and that estrogen status had no impact on total IgG sialylation on both Fab and Fc domains or general glycoprotein sialylation evaluated by ELISA. Furthermore, E2 status did not affect the total sialic acid content of total cells in lymphoid organs and neither B cells nor plasma cells. The study concluded that E2 treatment does not affect total serum glycoprotein sialylation but alters IgG glycosylation, including IgG sialylation, implying that estrogen functions as an intrinsic modulator of IgG sialylation and could thereby be one pathway by which estrogen modulates immunity.

Novel Insight into the Etiology of Haff Disease by Mapping the N-Glycome with Orthogonal Mass Spectrometry

Consumption of boiled crayfish may lead to Haff disease (HD), which is considered to result from an unidentified toxin, although the etiology is still obscure. Profiling of the N -glycome in HD would assist in deciphering the underlying molecular mechanism of the disease, whereas HD-associated glycosylation has never been explored. Herein, we enrolled 90 serum samples with HD patients and healthy controls from the Wuhan Center for Disease Control & Prevention between 2019 and 2020. N -glycome profiles of both serum and serum-derived immunoglobulin G (IgG) in HD were characterized by means of high-throughput-based orthogonal mass spectrometry. It was observed that HD is associated with an increase in the core fucosylation and mono-galactosylation of total serum glycoproteins. The serum level of IgG was found to serve as a good indicator for HD patients. In addition, differential galactosylation and sialylation of IgG were strongly correlated with HD. It was notable that the changes in the galactosylation and sialylation of IgG1 and IgG2 were subclass specific. Interestingly, altered sialylation and galactosylation of IgG2 or IgG3/4 strongly correlated with clinical markers for HD. Our study reveals the association of differential IgG N-glycosylation with HD, providing new insight into the etiology of this rare disease.

N-glycans from serum IgG and total serum glycoproteins specific for endometriosis

Endometriosis is a chronic inflammatory gynaecological disease characterized by the growth of endometrial tissue outside the uterine cavity. There are currently no definitive non-invasive diagnostic tools. Glycosylation is the most common posttranslational modification of proteins and altered glycosylation has been found in many diseases, including chronic inflammatory conditions and cancer. Sialylation and galactosylation on serum IgG have previously been found to be altered in endometriosis and serum sialylation changed after Zoladex (Goserelin Acetate) therapy. Using IgG and whole serum glycoproteins, we investigated N-glycosylation in two clinical cohorts of women with and without endometriosis. PNGase F-digested serum samples were fluorescently labelled and N-glycans were profiled by ultra-performance liquid chromatography. Clinical data was collected to link glycomic findings with metabolic and hormonal profiles. Total serum glycoprotein and IgG glycosylation differed in patients with endometriosis compared to control cases. The most significantly altered was glycan peak 3 from IgG, containing bisected biantennary glycans, which was decreased in the endometriosis cohorts (p = 0.0000005–0.018). In conclusion, this is the first pilot study to identify changes in N-glycans from whole serum glycoproteins associated with endometriosis. A larger validation study is now warranted and such studies should include the follow-up of surgically and pharmacologically treated patients.

P–340 Novel non-invasive diagnostic options for endometriosis - based on glycome analysis

Abstract Study question Could glycosylation changes on serum and/or urine glycoproteins be suitable biomarkers for the non-invasive diagnosis of endometriosis? Summary answer The glycosylation pattern on serum and urine glycoproteins differed significantly in endometriosis patients compared to controls, suggesting a novel role as biomarkers of the disease. What is known already There is little published on endometriosis and glycosylation, and most of the studies are conducted with tissue or peritoneal fluid samples, collected by invasive means. An Iraqi study draws attention to the importance of serum sialylation, which is dramatically changed in endometriosis patients after zoladex therapy, indicating that changes in serum sialylation may be a new biomarker of the disease. While glycosylation of urine in endometriosis has not been studied so far, in a study of endometrial cancer, the urinary level of two glycoproteins was significantly increased in the patients compared to the control group. Study design, size, duration This was a prospective study. In this basic research project, serum and urine samples were collected for glycome analysis in women with and without endometriosis, as diagnosed at laparoscopy. Glycated haemoglobin (HbA1c), fasting glucose levels as well as hormone levels were also collected from the patients to link our glycomic findings with metabolic and hormone profiles. The study was approved by the Research and Ethics Committee of the National Maternity Hospital, Dublin (EC19.2018). Participants/materials, setting, methods Samples from 24 cases of endometriosis (patients without previous anti-inflammatory or hormonal therapy, endometriosis was confirmed by laparoscopy) and 27 control patients (patients without endometriosis) were processed to analyse N-glycans (total serum), urine glycoproteins, and IgG. The pre-processed, PNGase F-digested serum and urine samples were labelled with fluorescent tag and then analysed by mass spectrometry, ultra-performance liquid chromatography (UPLC) in combination with exoglycosidase digestions and Glycostore (https://glycostore.org/). Main results and the role of chance Glycosylation on total serum and urine glycoproteins and IgG was investigated and differed in endometriosis compared to controls. The N-glycome from the total glycoproteins in serum and urine was also different. The proportion of the galactosylation and sialylation differed between urine and serum IgG and these alterations have an impact on the IgG function. Our preliminary data indicate, that there is an increase in alpha 2–3 sialylation, galactosylation, and fucosylation on urine glycans from endometriosis patients compared to the control pool. Urine is a good source of biomarkers as it can be collected non-invasively. Our group is the first to have developed a protocol for the recovery of N-glycans in urine and to have identified the total N-glycome in urine. The urine N-glycome contains mostly complex N-glycans and also some oligomannosylated and hybrid glycans. Our results may lead to non-invasive biomarkers for the diagnosis of endometriosis and the monitoring of the disease. Limitations, reasons for caution The number of participants involved in this basic research is low but this is a pilot study. A larger, validation study, is warranted in the future. Furthermore, the follow-up of treated patients also would be an interesting field of research. Wider implications of the findings: Glycomics may be a potent source of biomarkers of endometriosis, with a number of glyco-biomarkers already approved by the FDA. Endometriosis-associated glycomic profiles from serum and/or urine glycoproteins may represent viable targets for development of innovative non-invasive or minimally invasive diagnostics in this debilitating disease. Trial registration number Not applicable

HILIC-UPLC-MS for high throughput and isomeric N-glycan separation and characterization in Congenital Disorders Glycosylation and human diseases.

N-glycan analyses may serve uncovering disease-associated biomarkers, as well as for profiling distinctive changes supporting diagnosis of genetic disorders of glycan biosynthesis named congenital disorders of glycosylation (CDG). Strategies based on liquid chromatography (LC) preferentially coupled to electrospray ionization (ESI) - mass spectrometry (MS) have emerged as powerful analytical methods for N-glycan identification and characterization. To enhance detection sensitivity, glycans are commonly labelled with a functional tag prior to LC-MS analysis. Since most derivatization techniques are notoriously time-consuming, some commercial analytical kits have been developed to speed up N-deglycosylation and N-glycan labelling of glycoproteins of pharmaceutical and biological interest such as monoclonal antibodies (mAbs). We exploited the analytical capabilities of RapiFluor-MS (RFMS) to perform, by a slightly modified protocol, a detailed N-glycan characterization of total serum and single serum glycoproteins from specific patients with CDG (MAN1B1-CDG, ALG12-CDG, MOGS-CDG, TMEM199-CDG). This strategy, accomplished by Hydrophilic Interaction Chromatography (HILIC)-UPLC-ESI-MS separation of the RFMS derivatized N-glycans, allowed us to uncover structural details of patients serum released N-glycans, thus extending the current knowledge on glycan profiles in these individual glycosylation diseases. The applied methodology enabled to differentiate in some cases either structural isomers and isomers differing in the linkage type. All the here reported applications demonstrated that RFMS method, coupled to HILIC-UPLC-ESI-MS, represents a sensitive high throughput approach for serum N-glycome analysis and a valuable option for glycan detection and separation particularly for isomeric species.

N-Glycosylation of Serum IgG and Total Glycoproteins in MAN1B1 Deficiency.

MAN1B1-CDG has recently been characterized as a type II congenital disorder of glycosylation (CDG), disrupting not only protein N-glycosylation but also general Golgi morphology. Using our high-throughput, quantitative ultra-performance liquid chromatography assay, we achieved a detailed characterization of the glycosylation changes in both total serum glycoproteins and isolated serum IgG from ten previously reported MAN1B1-CDG patients. We have identified and quantified novel hybrid high-mannosylated MAN1B1-CDG-specific IgG glycans and found an increase of sialyl Lewis x (sLex) glycans on serum proteins of all patients. This increase in sLex has not been previously reported in any CDG. These findings may provide insight into the pathophysiology of this CDG.

High-Mannose Glycans are Elevated during Breast Cancer Progression

Alteration in glycosylation has been observed in cancer. However, monitoring glycosylation changes during breast cancer progression is difficult in humans. In this study, we used a well-characterized transplantable breast tumor mouse model, the mouse mammary tumor virus-polyoma middle T antigen, to observe early changes in glycosylation. We have previously used the said mouse model to look at O-linked glycosylation changes with breast cancer. In this glycan biomarker discovery study, we examined N-linked glycan variations during breast cancer progression of the mouse model but this time doubling the number of mice and blood draw points. N-glycans from total mouse serum glycoproteins were profiled using matrix-assisted laser desorption/ionization Fourier transform-ion cyclotron resonance mass spectrometry at the onset, progression, and removal of mammary tumors. We observed four N-linked glycans, m/z 1339.480 (Hex(3)HexNAc), 1485.530 (Hex(3)HexNAc(4)Fuc), 1809.639 (Hex(5)HexNAc(4)Fuc), and 1905.630 (Man(9)), change in intensity in the cancer group but not in the control group. In a separate study, N-glycans from total human serum glycoproteins of breast cancer patients and controls were also profiled. Analysis of human sera using an internal standard showed the alteration of the low-abundant high-mannose glycans, m/z 1419.475, 1581.528, 1743.581, 1905.634 (Man(6-9)), in breast cancer patients. A key observation was the elevation of a high-mannose type glycan containing nine mannoses, Man(9), m/z 1905.630 in both mouse and human sera in the presence of breast cancer, suggesting an incompletion of the glycosylation process that normally trims back Man(9) to produce complex and hybrid type oligosaccharides.

A strategy to reveal potential glycan markers from serum glycoproteins associated with breast cancer progression

Aberrant glycosylation on glycoproteins that are either presented on the surface or secreted by cancer cells is a potential source of disease biomarkers and provides insights into disease pathogenesis. N-Glycans of the total serum glycoproteins from advanced breast cancer patients and healthy individuals were sequenced by HPLC with fluorescence detection coupled with exoglycosidase digestions and mass spectrometry. We observed a significant increase in a trisialylated triantennary glycan containing alpha1,3-linked fucose which forms part of the sialyl Lewis x epitope. Following digestion of the total glycan pool with a combination of sialidase and beta-galactosidase, we segregated and quantified a digestion product, a monogalactosylated triantennary structure containing alpha1,3-linked fucose. We compared breast cancer patients and controls and detected a 2-fold increase in this glycan marker in patients. In 10 patients monitored longitudinally, we showed a positive correlation between this glycan marker and disease progression and also demonstrated its potential as a better indicator of metastasis compared to the currently used biomarkers, CA 15-3 and carcinoembryonic antigen (CEA). A pilot glycoproteomic study of advanced breast cancer serum highlighted acute-phase proteins alpha1-acid glycoprotein, alpha1-antichymotrypsin, and haptoglobin beta-chain as contributors to the increase in the glycan marker which, when quantified from each of these proteins, marked the onset of metastasis in advance of the CA 15-3 marker. These preliminary findings suggest that specific glycans and glycoforms of proteins may be candidates for improved markers in the monitoring of breast cancer progression.

Increased levels of galactose-deficient IgG in sera of HIV-1-infected individuals

The IgG from sera of patients with chronic inflammatory diseases of autoimmune character or some chronic microbial infections is frequently deficient in galactose on N-linked glycans. However, this phenomenon has not been investigated at length in human viral infections. To evaluate the glycosylation of serum IgG in HIV-1-positive patients. Psathyrella velutina lectin was used in enzyme-linked immunosorbent and Western blot assays to determine glycosylation. In addition, gas-liquid chromatography and mass spectrometry were utilized to confirm the galactose deficiency observed in the lectin-binding assays. HIV-1-infected individuals had significantly higher levels of galactose-deficient IgG than healthy controls. In fact, the galactose deficiency of the N-linked glycans observed in other diseases was even more profound in HIV-1 infection. This deficiency was primarily restricted to IgG when total serum glycoproteins were evaluated and IgG1 was the subclass most affected in all patients. Also, a significant increase in lectin binding was observed on IgG2 and IgG4 from HIV-1-positive females compared with HIV-1-negative females. Identification of deficient galactosylation of serum IgG from HIV-1-infected patients extended the spectrum of diseases in which this phenomenon has been observed. In addition, the results suggest yet another aspect of immune dysfunction as a result of HIV-1 infection.

Hydrophilic Affinity Isolation and MALDI Multiple-Stage Tandem Mass Spectrometry of Glycopeptides for Glycoproteomics

In glycoproteomics, key structural issues, protein identification, locations of glycosylation sites, and evaluation of the glycosylation site microheterogeneity should be easily evaluated in a large number of glycoproteins, while mass spectrometry (MS) provides substantial information about individual purified glycoproteins. Considering that structural issues are elucidated by studying glycopeptides and that the tandem MS of a tryptic peptide composed of several amino acid residues is enough for protein identification, construction of an MS-based method handling tryptic glycopeptides would be of considerable benefit in research. To this end, a simple and efficient method, utilizing hydrophilic binding of carbohydrate matrixes such as cellulose and Sepharose to oligosaccharides, was successfully applied to the isolation of tryptic glycopeptides. Both peptide and oligosaccharide structures were elucidated by multiple-stage tandem MS (MS(n)) of the ions generated by matrix-assisted laser desorption/ionization (MALDI), as follows. The MALDI ion trap mass spectrum of a tryptic glycopeptide mixture from N-linked glycoproteins was composed of the [M + H]+ ions of component glycopeptides. Collision-induced dissociation (CID) of the glycopeptide [M + H]+ ion generated saccharide-spaced peaks, with an interval of, for example, 146, 162, and 203 Da, and their fragment ions corresponding to the peptide and peptide + N-acetylglucosamine (GlcNAc) species in the MS2 spectrum. The saccharide-spaced ladder served to outline oligosaccharide structures, which were then selected as precursors for subsequent MS(n) analyses. The peptide or peptide + GlcNAc ions in the MS2 spectrum or the corresponding ions abundant in the MS1 spectrum were subjected to CID for determination of peptide sequences, to identify proteins and their glycosylation sites. The strategy, isolation of glycopeptides followed by MS(n) analysis, efficiently characterized the structures of beta2-glycoprotein I with four N-glycosylation sites and was applied to an analysis of total serum glycoproteins.

Detailed glycan analysis of serum glycoproteins of patients with congenital disorders of glycosylation indicates the specific defective glycan processing step and provides an insight into pathogenesis.

The fundamental importance of correct protein glycosylation is abundantly clear in a group of diseases known as congenital disorders of glycosylation (CDGs). In these diseases, many biological functions are compromised, giving rise to a wide range of severe clinical conditions. By performing detailed analyses of the total serum glycoproteins as well as isolated transferrin and IgG, we have directly correlated aberrant glycosylation with a faulty glycosylation processing step. In one patient the complete absence of complex type sugars was consistent with ablation of GlcNAcTase II activity. In another CDG type II patient, the identification of specific hybrid sugars suggested that the defective processing step was cell type-specific and involved the mannosidase III pathway. In each case, complementary serum proteome analyses revealed significant changes in some 31 glycoproteins, including components of the complement system. This biochemical approach to charting diseases that involve alterations in glycan processing provides a rapid indicator of the nature, severity, and cell type specificity of the suboptimal glycan processing steps; allows links to genetic mutations; indicates the expression levels of proteins; and gives insight into the pathways affected in the disease process.

High-mannose structure of apolipoprotein-B from low-density lipoproteins of human plasma.

Human plasma low-density lipoproteins were purified by flotation followed by gel filtration. The protein moiety of the lipoproteins, apolipoprotein-B, which was detected by polyacrylamide gel electrophoresis as the only protein component, contained 4.4% (by weight) carbohydrate. Glycopeptides liberated from apolipoprotein-B by pronase were fractionated by affinity chromatography on concanavalin-A--Sepharose. The results indicated that high-mannose glycopeptides interacting strongly with the lectin comprise about 37% of the total monosaccharides of apolipoprotein-B. Thus, as compared to the total serum glycoproteins having about 5% of their monosaccharides in high-mannose glycopeptides, low-density lipoproteins are relatively enriched in these structures amounting up to about 10% of the total high-mannose oligosaccharides in serum. The rest of the carbohydrates in low-density lipoproteins are suggested to be mainly biantennary acidic oligosaccharides interacting weakly with concanavalin A. The oligomannosidic chains from native low-density lipoproteins and isolated glycopeptides were released by digestion with endo-beta-N-acetylglucosaminidase H. Thin-layer chromatography of the released oligosaccharides indicated that apolipoprotein-B contains five different oligomannosidic structures varying in the number of the mannose residues from Man5GlcNAc to Man9GlcNAc. Separation of the per-O-benzoylated high-mannose oligosaccharides by high-pressure liquid chromatography revealed the same polymeric structures in a molar ratio (from Man5 to Man9) of 10:2:3:2:3. Apolipoprotein-B in low-density lipoproteins was calculated to contain five high-mannose chains in total. The different high-mannose oligosaccharides liberated by endo-beta-N-acetylglucosaminidase H were isolated with high-pressure liquid chromatography after reduction with NaBH4, and subjected to methylation analysis with gas-liquid chromatography--mass spectrometry. The data of these studies and the results of exoglycosidase treatment suggest the following structure for the main high-mannose oligosaccharide: (formula: see text) The higher polymeric structures are composed of chains in which the Man5GlcNAc structure is continued by one to four Man(alpha 1-2) residues.

Inflammation-induced increases in dolichol synthesis and hydroxymethylglutaryl-coenzyme a reductase activity in mouse liver are prevented by a high-cholesterol diet but not by fasting

The inflammatory response in mammals is characterized by the synthesis in the liver of several N-linked serum glycoproteins called acute-phase reactants. In C57BL/6J mice, turpentine-induced inflammation was accompanied by increases in 3-hydroxy-3-methylglutaryl-coenzyme A reductase activity, dolichol synthesis, and dolichyl phosphoryl mannose synthesis. Cholesterol feeding, but not fasting, prevented these inflammation-induced increases in reductase activity and dolichol synthesis. However, the rate of incorporation of [ 3H]mannose into total serum glycoproteins was not affected by the high-cholesterol diet, and this rate increased during acute inflammation in control and cholesterol-fed mice.

Effect of Vitamin A Deficiency on Glycosylation of Rat Serum α1-Macroglobulin

We previously reported that synthesis of the serum glycoprotein α1-macroglobulin (α1-M) was decreased in vitamin A-deficient rats. In a reexamination of this conclusion we have shown that the primary synthetic change, reflected by labeled glucosamine uptake, was an underglycosylation which initially has no effect on the glycoprotein’s concentration in serum. The severity of this lesion developed progressively and was independent of the glycosylation of total serum glycoprotein, at least up to 11 days after onset of the deficiency. Less glucosamine was demonstrated chemically in a α1-M purified from rats in the first week of deficiency; α1-M from more severely deficient rats showed significant depressions of mannose, galactose, and glucosamine.vitamin A serum glycoprotein α1-macroglobulin glucosamine

虚血性心疾患における血清糖蛋白質の研究

To study about the chemical components of serum glycoprotein (GP) in ischemic heart diseases (IHD), column chromatography of Sephadex G-100 and cellulose electrophoresis were performed on sera in 6 cases of angina pectoris (AP), 10 cases of acute myocardial infarction (AMI), 10 cases of old myocardial infarction (OMI) and 10 cases of Healthy person (PH).The results as follows1) Total serum GP is higher in AP, AMI and OMI than in that of HP.2) Serum GP was fractionated into Fraction I (Fr I) and Fraction II (Fr II) by column chromatography. The molecular size was in order of FrI<Fr II. The concentration of Fr I was higher in AP, AMI and OMI than in that of HP, but the concentration of Fr II was higher only in AMI than in that of HP.3) Fr I was separated into three fractions of α2, β-and γ-GP by electrophoresis.Fr II was separated into four fractions of α1, α2-β-and γ-GP by electrophoresis.4) In AP and OMI, α2-GP of Fr I, γ-GP of FrI and γ-GP of Fr II were higher than in that of HPIn AMI, α1-GP of Fr I, α1-GP of Fr II, β-GP of Fr II and γ-GP of Fr II were higher, and β-GP of Fr I and α2-GP of Fr II were lower than that of HP.5) The components of serum GP in IHD were different from those in HP.

Serum protein alterations in adjuvant-induced arthritis.

The effects of adjuvant-induced arthritis in the rat on serum protein alterations were investigated in a longitudinal study. Injection of an emulsion of the Wax D fraction of the tubercle bacillus in oil elicited pronounced elevations in the α1-globulin, α2-globulin, and seromucoid fractions, and the total serum glycoprotein within 24 h. During the acute stage of the disease, significant increases were observed in protein-bound hexose, and in the fibrinogen, seromucoid, α1-, α2-, and β-globulin fractions. A new protein, α2-AP (acute phase) globulin, appeared in the serum concomitant with decreased levels of total protein, albumin, and γ-globulin. Remission was characterized by a reversal of the above changes. Spontaneous renewal of the arthritic symptoms caused comparable but less marked serum changes similar to those observed in the various clinical phases of rheumatic disease in man. It was concluded that although the serum changes parallel to a certain extent the degree of pathologic activity, they are not specific, and reflect a common systemic response to inflammation.

The systemic response to inflammation

Various acute phase reactants (APR) have been studied in rats with different types of inflammation. These APR (serum albumin, serum globulins, plasma fibrinogens, serum alpha-2-glycoprotein [GP], total serum glycoproteins, serum mucoproteins) and neutrophils and lymphocytes reflect the onset, intensity, and duration of many types of experimental inflammation. The APR and cellular changes, like the inflammatory processes eliciting them, are stereotyped. Chemically induced, chronic immunologic, and delayed hypersensitivity-type acute inflammatory reactions cannot be distinguished by the APR. The most sensitive and quantitatively reliable index for the assessment of the “anti-inflammatory” action of the various steroidal and nonsteroidal drugs is the behaviour of the serum alpha-2-glycoproteins. This is also the most sensitive index of the “anti-inflammatory” action of adrenocortical steroids (or their absence, as shown in adrenalectomized rats). The various serum protein changes in rats with inflammation occur in response to greater sequestration rates at the sites of inflammation. Acute and chronic inflammation can be induced with a variety of substances affecting cell membranes (Filipin, digitonin, saponin, nystatin, d-α-tocopherol, vitamin A, and etruscomycin, to name but a few). These inflammatory reactions are inhibited in vivo by both steroidal and nonsteroidal “anti-inflammatory” drugs. We have visualized the process of inflammation as a wave of leaky membranes. The protective effects of nonsteroidal “anti-inflammatory” drugs on heat-induced and hypotonic erthrocyte lysis in vitro partially confirm the idea. The anti-arthritic effects of cytarabine (cytosine arabinoside, an immunosuppressant drug) also have been studied. Cytarabine has no systemic or local “anti-inflammatory” effect in a variety of experimental assay systems. It inhibits completely the onset of adjuvant arthritis and renders animals tolerant to further adjuvant inoculations. The APR cannot be used exclusively to distinguish between the different steroidal, non-steroidal, and immunosuppressant types of “anti-inflammatory” drugs. They are useful, however, in the overall assessment of the “anti-inflammatory” effects of drugs in all types of inflammation. The APR should be incorporated as standard procedures in laboratories “discovering” new “anti-inflammatory” drugs. Emphasis is placed on the APR as necessary requirements for the assessment of “anti-inflammatory” drugs because for example, Filipin-induced hind-paw edema can be inhibited in vivo by the i.p. administration of digitonin, another lytic agent which produces inflammation.

The effects of periodic challenge on the response of alpha-2-AP globulin and other acute-phase reactants of rat serum to tissue injury.

The effects of periodic challenge with turpentine on the concentration of the acute-phase reactants of serum were studied in young, adult, male Sprague–Dawley rats. The initial injection of the phlogogenic agent was followed by rapid and significant increases in the concentrations of plasma fibrinogen, total serum glycoprotein, serum complement, and the α2-AP globulin and seromucoid fractions. Variable response patterns were observed on subsequent challenges and in the intervals between. The conclusion was drawn that the acute-phase reactants of serum do not respond in a uniform manner to repeated challenge with a phlogogenic agent, and by inference their synthesis must be regulated by several different mechanisms.

THE SERUM GLYCOPROTEIN RESPONSE FOLLOWING X-IRRADIATION AND PRIMARY IMMUNIZATION IN THE RABBIT

Serial determinations of total serum glycoprotein, seromucoid hexose, total serum protein and antibody levels were carried out in four groups of adult, male albino rabbits subjected to the following experimental procedures: (a) 400 r whole body X-irradiation, (b) immunization with 10 mg I131BSA, (c) irradiation followed by antigen injection 24 hours later, and (d) bleeding controls. The dose of radiation employed suppressed the antibody response of (c). Significant increases in total glycoprotein and seromucoid hexose occurred 24 hours after exposure in the irradiated groups and persisted for 4 days. Pronounced decreases in total protein were observed in (a), (b), and (c). It was concluded that increases in serum glycoproteins were related primarily to tissue injury.

Serum Glycoprotein Changes in Pregnancy.

SummaryDuring pregnancy, total serum protein and serum albumin concentration decreased, while the α- and β-globulin protein fractions increased and the γ-globulin protein fraction remained at a rather constant level throughout pregnancy. Total serum glycoprotein, expressed as protein bound hexose, increased throughout the gestational period. This was accompanied by a rise in the levels of all glycoprotein fractions expressed as mg per 100 ml. Relative to the protein content of each fraction, the glycoprotein content of the albumin and γ-globulin fractions increased during pregnancy, while the α1-, α2- and β-fractions decreased in relative percentage of bound carbohydrate. During the early post partum period, albumin and γ-globulin glycoprotein decreased, while α1-, α2- and β-globulin glycoprotein increased. These results indicate that striking alterations of plasma protein metabolism occur in pregnancy.