Large-scale analysis of structural brain asymmetries during neurodevelopment: Associations with age and sex in 4265 children and adolescents.

Only a small number of studies have assessed structural differences between the two hemispheres during childhood and adolescence. However, the existing findings lack consistency or are restricted to a particular brain region, a specific brain feature, or a relatively narrow age range. Here, we investigated associations between brain asymmetry and age as well as sex in one of the largest pediatric samples to date (n = 4265), aged 1-18 years, scanned at 69 sites participating in the ENIGMA (Enhancing NeuroImaging Genetics through Meta-Analysis) consortium. Our study revealed that significant brain asymmetries already exist in childhood, but their magnitude and direction depend on the brain region examined and the morphometric measurement used (cortical volume or thickness, regional surface area, or subcortical volume). With respect to effects of age, some asymmetries became weaker over time while others became stronger; sometimes they even reversed direction. With respect to sex differences, the total number of regions exhibiting significant asymmetries was larger in females than in males, while the total number of measurements indicating significant asymmetries was larger in males (as we obtained more than one measurement per cortical region). The magnitude of the significant asymmetries was also greater in males. However, effect sizes for both age effects and sex differences were small. Taken together, these findings suggest that cerebral asymmetries are an inherent organizational pattern of the brain that manifests early in life. Overall, brain asymmetry appears to be relatively stable throughout childhood and adolescence, with some differential effects in males and females.

Inflammation, neurotrophism and oxidative stress and childhood psychopathology in a large community sample.

OBJECTIVE: To investigate the association between peripheral biomarkers and child psychopathology in a large community sample. METHOD: A total of 625 aged 6- to 13-year old subjects were recruited from a community school-based study. Psychopathology was assessed using the Child Behaviour Checklist (CBCL). Psychiatric diagnosis was evaluated using the Development and Well-Being Assessment. The following biomarkers were examined in peripheral blood: brain-derived neurotrophic factor, cytokines (IL-2, IL-4, IL-6, IL-10, IL-17, IFN-g, and TNF-α), chemokines (eotaxin/CCL11, IP-10, MCP-1), cytokine receptors (sTNFR1 and sTNFR2), and the oxidative stress marker TBARS. RESULTS: We found significant associations between sTNFR2, eotaxin/CCL11 and CBCL total score, as well as with specific dimensions of psychopathology. There were different patterns of association between these biomarkers and psychological and behavioural symptoms in children with and without a mental disorder. TBARS, IL-6 and MCP-1 were more specific to some clusters of symptoms in children with a psychiatric diagnosis. CONCLUSION: Our data support the potential use of biomarkers, especially those involved in immune-inflammatory pathways, in investigating neurodevelopmental psychopathology. Their association with different dimensions of symptoms might be of useful when analyzing illness severity and clusters of symptoms within specific disorders.

Cost-effectiveness of early intervention in psychosis in low- and middle-income countries: economic evaluation from São Paulo, Brazil.

AIMS: The effectiveness and cost-effectiveness of early intervention for psychosis (EIP) services are well established in high-income countries but not in low- and middle-income countries (LMICs). Despite the scarcity of local evidence, several EIP services have been implemented in LMICs. Local evaluations are warranted before adopting speciality models of care in LMICs. We aimed to estimate the cost-effectiveness of implementing EIP services in Brazil. METHODS: A model-based economic evaluation of EIP services was conducted from the Brazilian healthcare system perspective. A Markov model was developed using a cohort study conducted in São Paulo. Cost data were retrieved from local sources. The outcome of interest was the incremental cost-effectiveness ratio (ICER) measured as the incremental costs over the incremental quality-adjusted life-years (QALYs). Sensitivity analyses were performed to test the robustness of the results. RESULTS: The study included 357 participants (38% female), with a mean (SD) age of 26 (7.38) years. According to the model, implementing EIP services in Brazil would result in a mean incremental cost of 4,478 Brazilian reals (R$) and a mean incremental benefit of 0.29 QALYs. The resulting ICER of R$ 15,495 (US dollar [USD] 7,640 adjusted for purchase power parity [PPP]) per QALY can be considered cost-effective at a willingness-to-pay threshold of 1 Gross domestic product (GDP) per capita (R$ 18,254; USD 9,000 PPP adjusted). The model results were robust to sensitivity analyses. CONCLUSIONS: This study supports the economic advantages of implementing EIP services in Brazil. Although cultural adaptations are required, these data suggest EIP services might be cost-effective even in less-resourced countries.

Source memory errors in schizophrenia, hallucinations and negative symptoms: a synthesis of research findings.

BACKGROUND: Previous research has shown associations between source memory errors and hallucinations in patients with schizophrenia. We bring together here findings from a broad memory investigation to specify better the type of source memory failure that is associated with auditory and visual hallucinations. METHOD: Forty-one patients with schizophrenia and 43 healthy participants underwent a memory task involving recall and recognition of lists of words, recognition of pictures, memory for temporal and spatial context of presentation of the stimuli, and remembering whether target items were presented as words or pictures. RESULTS: False recognition of words and pictures was associated with hallucination scores. The extra-list intrusions in free recall were associated with verbal hallucinations whereas the intra-list intrusions were associated with a global hallucination score. Errors in discriminating the temporal context of word presentation and the spatial context of picture presentation were associated with auditory hallucinations. The tendency to remember verbal labels of items as pictures of these items was associated with visual hallucinations. Several memory errors were also inversely associated with affective flattening and anhedonia. CONCLUSIONS: Verbal and visual hallucinations are associated with confusion between internal verbal thoughts or internal visual images and perception. In addition, auditory hallucinations are associated with failure to process or remember the context of presentation of the events. Certain negative symptoms have an opposite effect on memory errors.

Dopamine transporter regulation during four nights of REM sleep deprivation followed by recovery--an in vivo molecular imaging study in humans.

OBJECTIVES: To assess the influence of total or selective REM sleep deprivation on the dopamine transporter (DAT) densities and sleep patterns of healthy volunteers. DESIGN: Prospective study. SETTING: Evaluation of polysomnography recordings and DAT density after 4 nights of selective REM sleep deprivation followed by 3 nights of sleep recovery compared to a control group and a group that was subjected to 2 nights of total sleep deprivation. Single positron emission computed tomography and [99mTc]TRODAT-1 were used to assess the cerebral DAT density in the striatum at baseline, after REM sleep deprivation and total sleep deprivation as well as after sleep recovery. Blood was collected daily to examine prolactin and estradiol levels, which were correlated with dopaminergic activity. PATIENTS OR PARTICIPANTS: Thirty healthy male volunteers ranging from 19 to 29 years of age were randomly assigned to one of three experimental groups after giving written informed consent (10 non-sleep deprived, 10 total sleep deprived, and 10 REM sleep deprived). MEASUREMENTS AND RESULTS: Four nights of REM sleep deprivation and 2 nights of total sleep deprivation induced distinct and heterogeneous patterns of sleep recovery. No significant modulation of DAT availability was observed within groups. In the recovery nights, changes in cortisol, prolactin and estradiol concentrations were significantly correlated with specific sleep stages in the total and REM sleep deprived groups. In addition, DAT density was positively correlated with estradiol concentration and inversely associated with SWS latency only after total sleep deprivation. CONCLUSION: Our study demonstrates that although sleep deprivation did not promote significant alterations in DAT density within the striatum, there were significant correlations among transporter availability, hormonal concentrations and sleep parameters.

Hallucinations and two types of free-recall intrusion in schizophrenia.

BACKGROUND: Previous research has demonstrated that various types of verbal source memory error are associated with positive symptoms in patients with schizophrenia. Notably, intrusions in free recall have been associated with hallucinations and delusions. We tested the hypothesis that extra-list intrusions, assumed to arise from poor monitoring of internally generated words, are associated with verbal hallucinations and that intra-list intrusions are associated with global hallucination scores. METHOD: A sample of 41 patients with schizophrenia was administered four lists of words, followed by free recall. The number of correctly recalled words and the number of extra- and intra-list intrusions were tallied. RESULTS: The verbal hallucination score was significantly correlated with the number of extra-list intrusions, whereas it was unrelated to the number of correctly recalled words. The number of intra-list intrusions was significantly correlated with the global, but not with the verbal, hallucination score in the subsample of hallucinating patients. It was marginally significantly correlated with the delusion score in delusional patients. CONCLUSIONS: The data corroborate the view that verbal hallucinations are linked to defective monitoring of internal speech, and that errors in context processing are involved in hallucinations and delusions.

Lower dopamine transporter density in an asymptomatic patient with Kleine-Levin syndrome.

BACKGROUND: Kleine-Levin syndrome (KLS) is a rare disorder whose pathophysiological mechanisms remain unknown. PATIENTS AND METHODS: To investigate dopamine abnormalities in KLS, a [99mTc]-TRODAT-1 single photon emission computerized tomography (SPECT) was performed in a patient with KLS during the asymptomatic period and compared with three matched healthy controls. RESULTS: The patient had 14% lower striatal dopamine transporter binding potential (DAT-BP) compared to the mean DAT-BP of three healthy controls. CONCLUSION: This study provides in vivo evidence for abnormalities in the DAT-BP, suggesting an involvement of the dopaminergic system in the pathophysiology of KLS.

Expression of the Tomato Pto Gene in Tobacco Enhances Resistance to Pseudomonas syringae pv tabaci Expressing avrPto.

The Pto gene encodes a serine-threonine kinase that confers resistance in tomato to Pseudomonas syringae pv tomato strains expressing the avirulence gene avrPto. We examined the ability of Pto to function in tobacco, a species that is sexually incompatible with tomato. Evidence that a heterologous Pto-like signal transduction pathway is present in tobacco was suggested by the fact that tobacco line Wisconsin-38 exhibits a hypersensitive response after infection with P. syringae pv tabaci expressing avrPto. We introduced a Pto transgene into cultivar Wisconsin-38 and assessed the ability of transformed plants to further inhibit growth of the P. s. tabaci strain expressing avrPto. The Pto-transformed tobacco plants exhibited a significant increase in resistance to the avirulent P. s. tabaci strain compared with wild-type tobacco as indicated by (1) more rapid development of a hypersensitive resistance response at high inoculum concentrations (108 colony-forming units per mL); (2) lessened severity of disease symptoms at moderate inoculum concentrations (106 and 107 colony-forming units per mL); and (3) reduced growth of avirulent P. s. tabaci in inoculated leaves. The results indicate that essential components of a Pto-mediated signal transduction pathway are conserved in tobacco and should prompt examination of resistance gene function across even broader taxonomic distances.

Regulation of the Osmotin Gene Promoter.

By introducing a chimeric gene fusion of the osmotin promoter and [beta]-glucuronidase into tobacco by Agrobacterium-mediated transformation, we have demonstrated a very specific pattern of temporal and spatial regulation of the osmotin promoter during normal plant development and after adaptation to NaCl. We have found that the osmotin promoter has a very high natural level of activity in mature pollen grains during anther dehiscence and in pericarp tissue at the final, desiccating stages of fruit development. GUS activity was rapidly lost after pollen germination. The osmotin promoter thus appears to be unique among active pollen promoters described to date in that it is active only in dehydrated pollen. The osmotin promoter was also active in corolla tissue at the onset of senescence. Adaptation of plants to NaCl highly stimulated osmotin promoter activity in epidermal and cortex parenchyma cells in the root elongation zone; in epidermis and xylem parenchyma cells in stem internodes; and in epidermis, mesophyll, and xylem parenchyma cells in developed leaves. The spatial and temporal expression pattern of the osmotin gene appears consistent with both osmotic and pathogen defense functions of the gene.

Plant Defense Genes Are Synergistically Induced by Ethylene and Methyl Jasmonate.

Combinations of ethylene and methyl jasmonate (E/MeJA) synergistically induced members of both groups 1 and 5 of the pathogenesis-related (PR) superfamily of defense genes. E/MeJA caused a synergistic induction of PR-1b and osmotin (PR-5) mRNA accumulation in tobacco seedlings. E/MeJA also synergistically activated the osmotin promoter fused to a [beta]-glucuronidase marker gene in a tissue-specific manner. The E/MeJA responsiveness of the osmotin promoter was localized on a -248 to +45 fragment that exhibited responsiveness to several other inducers. E/MeJA induction also resulted in osmotin protein accumulation to levels similar to those induced by osmotic stress. Of the several known inducers of the osmotin gene, including salicylic acid (SA), fungal infection is the only other condition known to cause substantial osmotin protein accumulation in Wisconsin 38, a tobacco cultivar that does not respond hypersensitively to tobacco mosaic virus. Based on the ability of the protein kinase C inhibitor 1-(5-isoquinolinylsulfonyl)-2-methylpiperazine to block ethylene induction of PR-1b mRNA accumulation and its inability to block osmotin mRNA induction by ethylene, these two PR gene groups appeared to have at least partially separate signal transduction pathways. Stimulation of osmotin mRNA accumulation by okadaic acid indicated that another protein kinase system is involved in regulation of the osmotin gene. SA, which is known to induce pathogen resistance in tobacco, could not induce the osmotin gene as much as E/MeJA and neither could it induce PR-1b as much as SA and MeJA combined.

Molecular imaging in hereditary forms of parkinsonism.

The development of in vivo molecular imaging to evaluate the dopamine (DA) system with positron-emission tomography and single photon emission computed tomography has been of key importance on monitoring in vivo nigrostriatal neuronal loss in Parkinson's disease (PD), mostly through assessments of pre- and post-synaptic DA receptors. The discoveries of genes related to hereditary forms of parkinsonism (PARK1, PARK2, PARK6, PARK7 and PARK8) have increased our understanding either of distinct subtypes of clinical expression in PD or its etiology. This article revises current data on molecular neuroimaging of genetic forms of parkinsonism comparing and contrasting its main features with the classical sporadic forms. Awareness of the spectrum variance in the genotype and its respective PD phenotype are useful to distinguish different pathophysiological mechanisms of PD.

Coordinated brain development: exploring the synchrony between changes in grey and white matter during childhood maturation.

Brain development during childhood and early adolescence is characterized by global changes in brain architecture. Neuroimaging studies have revealed overall decreases in cortical thickness (CT) and increases in fractional anisotropy (FA). Furthermore, previous studies have shown that certain cortical regions display coordinated growth during development. However, there is significant heterogeneity in the timing and speed of these developmental transformations, and it is still unclear whether white and grey matter changes are co-localized. In this multimodal neuroimaging study, we investigated the relationship between grey and white matter developmental changes and asynchronous maturation within brain regions in 249 normally developing children between the ages 7-14. We used structural magnetic resonance imaging (MRI) and diffusion tensor imaging (DTI) to analyze CT and FA, respectively, as well as their covariance across development. Consistent with previous studies, we observed overall cortical thinning with age, which was accompanied by increased FA. We then compared the coordinated development of grey and white matter as indexed by covariance measures. Covariance between grey matter regions and the microstructure of white matter tracts connecting those regions were highly similar, suggesting that coordinated changes in the cortex were mirrored by coordinated changes in their respective tracts. Examining within-brain divergent trajectories, we found significant structural decoupling (decreased covariance) between several brain regions and tracts in the 9- to 11-year-old group, particularly involving the forceps minor and the regions that it connects to. We argue that this decoupling could reflect a developmental pattern within the prefrontal region in 9- and 11-year-old children, possibly related to the significant changes in cognitive control observed at this age.

Gene expression alterations related to mania and psychosis in peripheral blood of patients with a first episode of psychosis.

Psychotic disorders affect ~3% of the general population and are among the most severe forms of mental diseases. In early stages of psychosis, clinical aspects may be difficult to distinguish from one another. Undifferentiated psychopathology at the first-episode of psychosis (FEP) highlights the need for biomarkers that can improve and refine differential diagnosis. We investigated gene expression differences between patients with FEP-schizophrenia spectrum (SCZ; N=53) or FEP-Mania (BD; N=16) and healthy controls (N=73). We also verified whether gene expression was correlated to severity of psychotic, manic, depressive symptoms and/or functional impairment. All participants were antipsychotic-naive. After the psychiatric interview, blood samples were collected and the expression of 12 psychotic-disorder-related genes was evaluated by quantitative PCR. AKT1 and DICER1 expression levels were higher in BD patients compared with that in SCZ patients and healthy controls, suggesting that expression of these genes is associated more specifically to manic features. Furthermore, MBP and NDEL1 expression levels were higher in SCZ and BD patients than in healthy controls, indicating that these genes are psychosis related (independent of diagnosis). No correlation was found between gene expression and severity of symptoms or functional impairment. Our findings suggest that genes related to neurodevelopment are altered in psychotic disorders, and some might support the differential diagnosis between schizophrenia and bipolar disorder, with a potential impact on the treatment of these disorders.

Effect of nitrogen starvation on the level of adenosine 3',5'-monophosphate in Anabaena variabilis.

Low levels of adenosine 3',5'-monophosphate (cyclic AMP) were detected in the cyanobacterium Anabaena variabilis using a protein binding assay and two radioisotopic labelling methods. The basal concentration of intracellular cyclic AMP ranged from 0.27 pmol/mg protein in A. variabilis Kutz grown under heterotrophic conditions to 1.0--2.7 pmol/mg protein in A. variabilis strain 377 grown autotrophically. Extracellular cyclic AMP was found to comprise as much as 90% of the total cyclic AMP in rapidly growing cultures. When A. variabilis strain 377 was starved of nitrogen, a 3--4-fold increase in intracellular cyclic AMP was observed during the 24 h period coincident with early heterocyst development.

NaCl-Induced Alterations in Both Cell Structure and Tissue-Specific Plasma Membrane H+ -ATPase Gene Expression.

NaCl-induced plasma membrane H+-ATPase gene expression, which occurs in roots and fully expanded leaves of the halophyte Atriplex nummularia L. (X. Niu, M.L. Narasimhan, R.A. Salzman, R.A. Bressan, P.M. Hasegawa [1993] Plant Physiol 103: 713-718), has been differentially localized to specific tissues using in situ RNA hybridization techniques. Twenty-four-hour exposure of plants to 400 mM NaCl resulted in substantial accumulation of H+ pump message in the epidermis of the root tip and the endodermis of the root elongation/differentiation zone. In expanded leaves, NaCl induction of plasma membrane H+-ATPase message accumulation was localized to bundle-sheath cells. Ultrastructural analyses indicated that significant cytological adaptations in root cells included plasmolysis that is accompanied by plasma membrane invaginations, formation of Hechtian strands and vesiculation, and vacuolation. These results identify specific tissues that are involved in the regulation of Na+ and Cl- uptake into different organs of the halophyte A. nummularia and provide evidence of the intercellular and interorgan coordination that occurs in the mediation of NaCl adaptation.

Salt-Sensitive Mutants of Chlamydomonas reinhardtii Isolated after Insertional Tagging.

We describe the isolation of salt-sensitive Chlamydomonas reinhardtii mutants by insertional mutagenesis using the nitrate reductase (Nit1) gene. The plasmid pMN24, containing Nit1, was used for transformation of 305CW15 (nit1 cw15 mt+), and transformants were selected for complementation of the nit- phenotype. From 6875 nit+ colonies, four transformants (S4, S18, S46, and S66) were isolated that exhibited both Na+ and Li+ sensitivity (sod-), and another transformant (S33) was selected that exhibited sensitivity to Li+ but not Na+ (lit-) based on relative growth comparisons with the wild-type strain. S33, S46, and S66 were no more growth inhibited by sorbitol than was 305CW15. In comparison, S4 and S18 exhibited substantial growth inhibition in medium supplemented with sorbitol. Genetic analyses indicated that the salt-sensitive mutants were each defective in a single recessive gene. The mutant genes in S4 (sod1), S33 (lit1), and S66 (sod3) are linked to a functional copy of Nit1 and are presumably tagged with a pMN24 insertion.

The modest but growing Brazilian presence in psychiatric, psychobiological and mental health research: assessment of the 1998-2002 period.

The objective of the present survey was to assess the Brazilian scientific production in psychiatry, psychobiology, and mental health during the 1998-2002 period. The universities' graduate programs concentrate the vast majority of the scientific production in Brazil. We assessed the annual reports from the graduate programs to the Brazilian Ministry of Education concerning master's and doctoral theses and the articles published in journals indexed by the Institute of Scientific Information (ISI). There are nine Master's and Doctoral graduate programs dedicated to research in psychiatry, neuropsychiatry, psychobiology, and mental health in the country, seven being located in southern states. During the 5-year period, from 1998 to 2002, 186 students received their doctorate degree (37/year). The programs published 637 articles in journals indexed by ISI, the majority of them in journals with an impact factor higher than 2. The research advisors' productivity varied among graduate programs, ranging from 0.6 to 2.0 articles per year in ISI-indexed journals. Despite the substantial barriers faced by the Brazilian scientific community (mainly financial and writing difficulties), Brazil's scientific mental health production is on the rise. The number of articles published in ISI-indexed journals has doubled without a significant increase in the number of graduate theses, suggesting that there was an improvement in both the quality of the scientific production and the productivity of the graduate programs. Based on these data, it is reasonable to predict a tendency to an increase in production over the next few years.

Revictimization of violence suffered by those diagnosed with alcohol dependence in the general population.

OBJECTIVE: To verify the association between violence and alcohol dependence syndrome in sample populations. METHOD: Population-wide survey with multistage probabilistic sample. 3,744 individuals of both genders, aged from 15 to 75 years, were interviewed from the cities of São Paulo and Rio de Janeiro using the Composite International Diagnostic Interview (CIDI 2.1). RESULTS: In both cities, alcohol dependence was associated with the male gender, having suffered violence related to criminality, and having suffered familial violence. In both cities, urban violence, in more than 50% of cases, and familial violence, in more than 90% of cases, preceded alcohol dependence. The reoccurrence of traumatic events occurred in more than half of individuals dependent on alcohol. In São Paulo, having been diagnosed with PTSD is associated with violence revictimization (P = 0.014; Odds = 3.33). CONCLUSION: Alcohol dependence syndrome is complexly related to urban and familial violence in the general population. Violence frequently precedes alcoholism, but this relationship is dependent on residence and traumatic events. This vicious cycle contributes to perpetuating the high rates of alcoholism and violence in the cities. Politicians ordering the reduction of violence in the large metropolises can, potentially, reduce alcoholism and contribute to the break of this cycle.

Convergent evidences from human and animal studies implicate angiotensin I-converting enzyme activity in cognitive performance in schizophrenia.

In schizophrenia (SCZ), higher angiotensin I-converting enzyme (ACE) levels have been reported in patient's blood and cerebrospinal fluid (CSF). Hereby, we propose to explore whether the ACE activity levels are associated to cognitive performance in SCZ. Seventy-two patients with SCZ or schizoaffective disorder diagnosis, and 69 healthy controls (HCs) underwent a cognitive battery with parallel collection of peripheral blood samples to measure ACE activity. Significant higher ACE activity levels were confirmed in the plasma of SCZ patients compared with HCs (Student's t=-5.216; P<0.001). ACE activity significantly correlated to Hopkins delayed recall measures (r=-0.247; P=0.004) and Hopkins total (r=-0.214; P=0.012). Subjects grouped as high ACE activity (above average) had worse performance compared with low ACE activity level group for Hopkins delayed recall measure, even after correction for clinical condition, age, gender and years of education (P=0.029). The adjusted R squared for this final model was 0.343. This result was evident only comparing extreme groups for ACE activity, when splitting the sample in three groups with similar number of subjects. To clarify this finding, we performed an evaluation of the cognitive performance of transgenic mice with three copies of ACE gene in novel object recognition (NOR) test, which showed that such animals presented impairment in NOR (P<0.05) compared with two copies of wild-type animals. The results observed in SCZ patients and animal model suggest both the association of ACE to cognitive deficits in SCZ. This finding may support the evaluation of novel treatment protocols and/or of innovative drugs for specific intervention of cognitive deficits in SCZ envisioning concomitant ACE activity and behavior evaluations.

Identification of N-acetylglucosamine binding residues in Griffonia simplicifolia lectin II.

Primary structure and crystallographic data of several legume lectins were used to predict the involvement in carbohydrate binding of six amino acid residues (Asp88, Glu108, Tyr134, Asn136, Leu226 and Gln227) in Griffonia simplicifolia lectin II (GS-II). The functional involvement of these residues was evaluated by assessing GlcNAc binding of modified forms of GS-II in which these residues were eliminated in truncated peptides or systematically substituted with other amino acids by site-specific mutations. Mutations at Asp88, Tyr134 or Asn136 eliminated GlcNAc binding activity by GS-II, while those at Glu108, Leu226 or Gln227 did not alter the activity. The former three amino acids were functionally essential for carbohydrate binding by GS-II presumably through hydrogen bonding to and hydrophobic interactions with GlcNAc. Although an Asp or Gly substitution for Tyr134 eliminated GlcNAc affinity, substitution with Phe did not appreciably affect binding. Despite the fact that mutations to Leu226 and Gln227 did not alter carbohydrate binding, a truncated form of GS-II lacking these residues no longer exhibited carbohydrate binding affinity.