Trans- and Multigenerational Maternal Social Isolation Stress Programs the Blood Plasma Metabolome in the F3 Generation.

Metabolic risk factors are among the most common causes of noncommunicable diseases, and stress critically contributes to metabolic risk. In particular, social isolation during pregnancy may represent a salient stressor that affects offspring metabolic health, with potentially adverse consequences for future generations. Here, we used proton nuclear magnetic resonance (1H NMR) spectroscopy to analyze the blood plasma metabolomes of the third filial (F3) generation of rats born to lineages that experienced either transgenerational or multigenerational maternal social isolation stress. We show that maternal social isolation induces distinct and robust metabolic profiles in the blood plasma of adult F3 offspring, which are characterized by critical switches in energy metabolism, such as upregulated formate and creatine phosphate metabolisms and downregulated glucose metabolism. Both trans- and multigenerational stress altered plasma metabolomic profiles in adult offspring when compared to controls. Social isolation stress increasingly affected pathways involved in energy metabolism and protein biosynthesis, particularly in branched-chain amino acid synthesis, the tricarboxylic acid cycle (lactate, citrate), muscle performance (alanine, creatine phosphate), and immunoregulation (serine, threonine). Levels of creatine phosphate, leucine, and isoleucine were associated with changes in anxiety-like behaviours in open field exploration. The findings reveal the metabolic underpinnings of epigenetically heritable diseases and suggest that even remote maternal social stress may become a risk factor for metabolic diseases, such as diabetes, and adverse mental health outcomes. Metabolomic signatures of transgenerational stress may aid in the risk prediction and early diagnosis of non-communicable diseases in precision medicine approaches.

Social Isolation Stress Modulates Pregnancy Outcomes and the Inflammatory Profile of Rat Uterus.

Prenatal stressors have been linked to adverse pregnancy outcomes; including preterm birth (PTB). Recent work demonstrates that social isolation in mothers represents a silent stressor contributing to PTB risk. Here; we investigate the association of inflammatory and stress markers with PTB risk in Long-Evans rats exposed to social isolation stress (SIS) during preconception and pregnancy across four generations (F0-F3). Gestational length; blood glucose; corticosterone levels; and maternal and offspring weights were assessed in two SIS paradigms: transgenerational (TG) and multigenerational (MG) exposure. Maternal uterine tissues were collected 21 days after the dams gave birth. Exposure to SIS reduced pregnancy lengths in the parental generation and neonatal birth weights in the F1 and F2 generations. Interleukin (IL)-1ß (Il1b) mRNA levels increased in F0 animals but decreased in the offspring of both stress lineages. Protein levels of IL-1ß decreased in the TG lineage. Corticotrophin-releasing hormone receptor 1 (Crhr1) expression decreased in SIS-exposed F0 animals and increased in the TG-F2 and MG-F1 offspring. Expression of enzyme 11-ß hydroxysteroid dehydrogenase-2 (11bHSD2) was enhanced in F1 animals. These findings suggest SIS has adverse consequences on the F0 mothers; but their F1-F3 progeny may adapt to this chronic stress; thus supporting the fetal programming hypothesis.

Multiple prenatal stresses increase sexual dimorphism in adult offspring behavior.

INTRODUCTION: Maternal gestational stress and immune activation have independently been associated with affective and neurodevelopmental disorders across the lifespan. We investigated whether rats exposed to prenatal maternal stressors (PNMS) consisting of psychological stress, interleukin (IL)-1ß or both (two-hit stress) during critical developmental windows displayed a behavioral phenotype representative of these conditions. METHODS: Long-Evans dams were exposed to psychological stressors consisting of restraint stress and forced swimming from gestational day (GD)12 to 18 or to no stress (controls). From GD17 until day of delivery, these same animals were injected with saline or IL-1ß as a second hit and immune stressor (5 µg/day, intraperitoneally). The behavior of F1 offspring adults was tested on the open field test, elevated plus maze and affective exploration task on postnatal days (P)90, 100 and 110 respectively. RESULTS: The effects of PNMS differed depending on the specific testing environment and potentially the age at assessment, especially in female offspring. Both locomotion and anxiety-like behavioral measures were susceptible to PNMS effects. In females, psychological stress increased anxiety-like behavior, whereas IL-1ß had an opposite effect, inducing exploration and risk-taking behavior on the open field test and the elevated plus maze. When present, interactions between both stressors limited the anxiogenic effect of psychological stress on its own. In contrast, prenatal psychological stress increased anxiety-like behavior in adult males overall. A similar anxiogenic effect of IL-1ß was only found on the open field test while the Stress*IL-1ß interaction appeared to limit the effect of either alone. Contrarily, the PNMS effects on anxiety-like behavior on the affective exploration task were highly similar between both sexes. Analysis of males and females together revealed an additive effect of Stress and IL-1ß on the number of exits from the refuge, a measure of risk assessment and thus correlated with anxiety. CONCLUSION: PNMS affected offspring adult behavior in a sex-dependent manner. Effects on females were more variable, whereas psychological stress mostly induced anxiety-like behavior in males. These data highlight the sexual dimorphism in vulnerability to prenatal stressors. Maternal or stress-induced programming of the stress response and neuroinflammation may play an important role in mediating stress effects on offspring adult behavior.

The rat cumulative allostatic load measure (rCALM): a new translational assessment of the burden of stress.

Determinants of lifetime health are complex and emphasize the need for robust predictors of disease risk. Allostatic load (AL) has become a clinical framework to estimate the cumulative biological burden associated with chronic stress. To assist knowledge translation in the developmental origins of health and disease field, clinically valid methods for reliable AL assessment in experimental models are urgently needed. Here, we introduce the rat cumulative allostatic load measure (rCALM), as a new preclinical knowledge translation tool to assess the burden of chronic stress. First, we identified an array of stress-associated physiological markers that are particularly sensitive to hypothalamic-pituitary-adrenal axis dysregulation by ancestral prenatal stress. Second, we determined which of these markers are susceptible to an intervention by environmental enrichment (EE) to mitigate AL. The markers most responsive to stress and EE therapy were assembled to become operationalized in the rCALM. Third, the new rCALM was validated for the ability to indicate future disease risks. The results show that the rCALM estimates the burden of chronic stress and serves as a proxy to estimate stress resilience and vulnerability to disease. Using the rCALM we showed that enrichment therapy can offset the adverse health outcomes linked to a high AL. Thus, the rCALM provides a model for the development of new test strategies that facilitate knowledge translation in preclinical animal models.

Prenatal two-hit stress affects maternal and offspring pregnancy outcomes and uterine gene expression in rats: match or mismatch?

Maternal stress and inflammation excesses can lead to adverse pregnancy outcomes and offspring development. We evaluated whether distinct prenatal stressors affect pregnancy, maternal and offspring outcomes, and uterine gene expression differently when combined than either alone. Long-Evans dams were exposed to psychological or/and (two-hit) immune stress (interleukin-1 beta [IL-1ß]), on gestational days 12-18 and 17-delivery, respectively. Gestational length, maternal weight gain, glycaemia and corticosterone levels, offspring weight, and gender effects were recorded. Maternal and offspring uteri were collected at weaning and on postnatal day 160 correspondingly. Uterine expression of genes involved in local progesterone metabolism, neuroendocrine and immune systems were analyzed using quantitative real-time polymerase chain reaction. Maternal two-hit stress increased gestational length variation and the occurrence of adverse pregnancy outcomes while reducing gestational weight gain. Pup weight was negatively affected by prenatal stressors in a gender-specific way. In dams, IL-1ß upregulated gene expression of neuroendocrine (Crh, Crhr1) and cytokine genes (Il1b, Il1rn, Il6, and Il10). Conversely, transcriptional patterns in offspring uteri were more variable with gene-specific up- or downregulation by each stressor separately, while exposure to both extensively reduced the expression of neuroendocrine (Hsd11b1), cytokine (Il1a, Il1rn, Il6), and IL-1 receptor genes. In conclusion, maternal stress affects physiological and molecular processes in dams and their offspring; two hits have different effects than single stressors. Outcomes appear generation-, gender-, and stressor-specific. Dampening of offspring uterine gene expression after exposure to multiple stressors could fit within the match/mismatch hypothesis of perinatal programming, with offspring preparing for a stressful life.

Upper Limb Intention Tremor in Multiple Sclerosis: An Evidence-Based Review of Assessment and Treatment.

BACKGROUND: To present the current knowledge on the characteristics, assessment, and treatment of upper limb intention tremor to inform and improve future intervention studies in patients with multiple sclerosis (MS), we conducted a literature review for articles on upper limb intention tremor in patients with MS. METHODS: Two reviewers conducted searches in PubMed, Web of Science, and MEDLINE (Ovid). Relevant articles, sorted on inclusion criteria, were examined for descriptions and assessments of upper limb intention tremor, and intervention studies were evaluated based on treatment type. RESULTS: Eight descriptive studies were found reporting on the incidence and severity of tremor, impairments, and lesion load. Ten studies focused on measurement of tremor using various assessments. Intervention studies included eight articles using a diverse set of noninvasive techniques mainly showing transient reduction in tremor amplitude and temporary increase in function. Eighteen studies on pharmacologic interventions were found, with most displaying positive outcomes and mediation of tremor; others showed little to no benefit. Surgical interventions included 17 studies on thalamotomy and 20 on deep brain stimulation. Most studies showed tremor improvement after surgery; however, most sample sizes were small, and interventions were highly invasive, with potential adverse effects resulting from surgery. CONCLUSIONS: The literature on upper limb intention tremor in MS is relatively sparse. More studies are required to determine mechanism of action and to provide more suitable and sustainable interventions to decrease upper limb intention tremor and improve quality of life of individuals with MS.

Environmental Intervention as a Therapy for Adverse Programming by Ancestral Stress.

Ancestral stress can program stress sensitivity and health trajectories across multiple generations. While ancestral stress is uncontrollable to the filial generations, it is critical to identify therapies that overcome transgenerational programming. Here we report that prenatal stress in rats generates a transgenerationally heritable endocrine and epigenetic footprint and elevated stress sensitivity which can be alleviated by beneficial experiences in later life. Ancestral stress led to downregulated glucocorticoid receptor and prefrontal cortex neuronal densities along with precocious development of anxiety-like behaviours. Environmental enrichment (EE) during adolescence mitigated endocrine and neuronal markers of stress and improved miR-182 expression linked to brain-derived neurotrophic factor (BDNF) and neurotrophin-3 (NT-3) regulation in stressed lineages. Thus, EE may serve as a powerful intervention for adverse transgenerational programming through microRNA-mediated regulation of BDNF and NT-3 pathways. The identification of microRNAs that mediate the actions of EE highlights new therapeutic strategies for mental health conditions and psychiatric disease.

Environmental enrichment mitigates the impact of ancestral stress on motor skill and corticospinal tract plasticity.

BACKGROUND: An adverse fetal environment in utero has been associated with long-term alterations in brain structure and function, and a higher risk of neurological disorders in later life. A common consequence of early adverse experience is impaired motor system function. A causal relationship for stress-associated impairments and a suitable therapy, however, have not been determined yet. OBJECTIVE: To investigate the impact of ancestral stress on corticospinal tract (CST) morphology and fine motor performance in rats, and to determine if adverse programming by ancestral stress can be mitigated by environmental enrichment therapy in rats. METHODS: The study examined F3 offspring generated by three lineages; one with prenatal stress only in the F1 generation, one with compounding effects of multigenerational prenatal stress, and a non-stress control lineage. F3 offspring from each lineage were injected with biotinylated dextran amine (BDA) into the motor cortex for anterograde tracing of the CST. RESULTS: Examination of the CST revealed reduced axonal density in the ancestrally stressed lineages. These anatomical changes were associated with significant impairments in skilled walking, as indicated by reduced foot placement accuracy and disturbed inter-limb coordination. Therapeutic intervention by environmental enrichment reduced the neuromorphological consequences of ancestral stress and restored skilled walking ability. CONCLUSIONS: The data suggest a causal relationship between stress-induced abnormal CST function and loss of fine motor performance. Thus, ancestral stress may be a determinant of motor system development and motor skill. Environmental enrichment may represent an effective intervention for the adverse programming by ancestral stress and trauma.

Altered brain morphology and functional connectivity reflect a vulnerable affective state after cumulative multigenerational stress in rats.

Prenatal stress is a risk factor for abnormal neuroanatomical, cognitive, behavioral and mental health outcomes with potentially transgenerational consequences. Females in general seem more resilient to the effects of prenatal stress than males. Here, we examined if repeated stress across generations may diminish stress resiliency and cumulatively enhance the susceptibility for adverse health outcomes in females. Pregnant female rats of three successive generations were exposed to stress from gestational days 12-18 to generate multigenerational prenatal stress (MPS) in the maternal lineage. Stress response was measured by plasma corticosterone levels and open-field exploration in each generation. Neuromorphological consequences of MPS were investigated in the F3 generation using in vivo manganese-enhanced magnetic resonance imaging (MEMRI), T2-relaxometry, and cytoarchitectonics in relation to candidate gene expression involved in brain plasticity and mental health. Each additional generation of prenatal stress incrementally elevated hypothalamic-pituitary-adrenal axis activation, anxiety-like and aversive behaviors in adult female offspring. Elevated stress responses in the MPS F3 generation were accompanied by reduced neural density in prefrontal cortex, hippocampus and whole brain along with altered brain activation patterns in in vivo MEMRI. MPS increased ephrin receptor A5 (Epha5), neuronal growth regulator (Negr1) and synaptosomal-associated protein 25 (Snap25) gene expression and reduced fibroblast growth factor 12 (Fgf12) in prefrontal cortex. These genes regulate neuronal maturation, arborization and synaptic plasticity and may explain altered brain cytoarchitectonics and connectivity. These findings emphasize that recurrent stress across generations may cumulatively increase stress vulnerability and the risk of adverse health outcomes through perinatal programing in females.

Environmental enrichment as an intervention for adverse health outcomes of prenatal stress.

Prenatal stress (PS) has complex neurological, behavioural and physiological consequences for the developing offspring. The phenotype linked to PS usually lasts into adulthood and may even propagate to subsequent generations. The often uncontrollable exposure to maternal stress and the lasting consequences emphasize the urgent need for treatment strategies that effectively reverse stress programming. Exposure to complex beneficial experiences, such as environmental enrichment (EE), is one of the most powerful therapies to promote neuroplasticity and behavioural performance at any time in life. A small number of studies have previously used EE to postnatally treat consequences of PS in the attempt to reverse deficits that were primarily induced in utero . This review discusses the available data on postnatal EE exposure in prenatally stressed individuals. The goal is to determine if EE is a suitable treatment option that reverses adverse consequences of stress programming and enhances stress resiliency. Moreover, this review discusses data with respect to relevant hypotheses including the cumulative stress and the mismatch hypotheses. The articles included in this review emphasize that EE reverses most behavioural, physiological and neural deficits associated with PS. Differing responses may be dependent on the timing and variability of stress and EE, exercise, and potentially vulnerable and resilient phenotypes of PS. Results from this study suggest that enrichment may provide an effective therapy for clinical populations suffering from the effects of PS or early life trauma.

Allostatic Load and Preterm Birth.

Preterm birth is a universal health problem that is one of the largest unmet medical needs contributing to the global burden of disease. Adding to its complexity is that there are no means to predict who is at risk when pregnancy begins or when women will actually deliver. Until these problems are addressed, there will be no interventions to reduce the risk because those who should be treated will not be known. Considerable evidence now exists that chronic life, generational or accumulated stress is a risk factor for preterm delivery in animal models and in women. This wear and tear on the body and mind is called allostatic load. This review explores the evidence that chronic stress contributes to preterm birth and other adverse pregnancy outcomes in animal and human studies. It explores how allostatic load can be used to, firstly, model stress and preterm birth in animal models and, secondly, how it can be used to develop a predictive model to assess relative risk among women in early pregnancy. Once care providers know who is in the highest risk group, interventions can be developed and applied to mitigate their risk.

The diagnosis of onychomycosis in a geriatric population: a study of 450 cases in South Florida.

An investigative study was performed to determine the diagnosis of onychomycosis in a South Florida geriatric population. In this study, 450 cases of suspected onychomycosis involving men and women 65 years of age and older from a private practice office and two nursing home settings were used. Samples were taken from the hallux toenail and sent to a mycology laboratory for fluorescent potassium hydroxide (KOH) preparation and microscopic examination of a fungal culture. Of the 450 cases studied, 46.4% of the patients had a single fungal organism cultured, 30.4% had a mixed fungal infection cultured, and 23.1% had no fungal growth. Saprophytes were found in 59.9% of the 526 total fungal organisms cultured while dermatophytes were found in only 23.8%. The results of this investigation demonstrate that there may be a shift from isolated dermatophyte infection to mixed saprophyte infections in a geriatric population with onychomycosis.

A "new" low-incidence red cell antigen, LOCR, associated with altered expression of Rh antigens.

BACKGROUND: Mild cases of hemolytic disease of the newborn were being studied when it was recognized that the propositi's red cells carried a novel antigen. STUDY DESIGN AND METHODS: Serologic and genetic studies of a new low-incidence antigen, LOCR (International Society of Blood Transfusion series number 700.53), were performed. RESULTS: The antigen is associated with altered expression of the Rh antigen c in two unrelated families and in a third proposita and with an altered expression of e in a fourth proposita. The lods for the gene controlling LOCR relative to Rh are 2.107 at theta = 0.00; that is, they fall short of the requirement for inclusion of the antigen in the Rh blood group system. LOCR is excluded from the ABO, MNS, Lutheran, Kell, Duffy, Kidd, Xg, Chido/Rodgers, Kx, and Gerbich blood group systems. Genetic evidence suggests it is not part of the Yt, Colton, or LW systems. CONCLUSION: Serologic evidence that LOCR is associated with altered expression of c or e strongly suggests that LOCR is part of the Rh blood group system. However, the genetic evidence falls short of the level required for system assignment.

Gene conversion confined to a direct repeat of the acceptor splice site generates allelic diversity at human glycophorin (GYP) locus.

The glycophorin locus (GYP) on the long arm of chromosome 4 encodes antigens of the MNSs blood group system and displays considerable allelic variation among human populations. The genomic structure and organization of a variant glycophorin allele specifying a novel Miltenberger (Mi)-related phenotype, MiX, were examined. This variant probably arose from a gene conversion event involving a direct repeat of the acceptor splice site. Southern blot analysis indicated that MiX gene derived its 5' and 3' portions from glycophorin B or delta gene but its internal part from glycophorin A or alpha gene. Genomic sequences encompassing the rearranged regions of the MiX gene were amplified by single copy polymerase chain reaction. Direct DNA sequencing showed that during the formation of MiX gene, a short stretch of alpha exon III with a donor splice site has replaced a silent sequence in the delta gene containing a cryptic acceptor splice site. The upstream delta-alpha breakpoint is flanked by the direct repeats of the acceptor splice site, whereas the down-stream alpha-delta breakpoint is located in the adjacent intron. This segmental transfer produced a new composite exon whose expression not only transactivated a portion of silent sequence but also created intraexon and interexon hybrid junctions that characterize the antigenic specificities of MiX glycophorin. The identification of MiX as yet another delta-alpha-delta hybrid different from MiIII and MiVI in gene conversion sites suggests that shuffling of expressed and unexpressed sequences through particular genomic DNA motifs has been an important mechanism for shaping the antigenic diversity of MNSs blood group system during evolution.

Typing of MNSs blood group specific sequences in the human genome and characterization of a restriction fragment tightly linked to S-s- alleles.

Human erythrocyte membrane alpha and delta glycophorins (glycophorins A and B) carry the antigens for the M,N,S,s blood group system. Synthetic oligonucleotides spanning coding regions for M,N,S, and s epitopes were used to examine DNAs from 50 individuals selected at random and from individuals known to exhibit S(-)s(-)U- or S(-)s(-)U+ blood group phenotypes. We showed that M,N,S,s, blood group-specific sequences occur as multicopies in the human genome and reside within the alpha and delta glycophorin genes and also within the third glycophorin gene (glycophorin E gene). DNA typing with M- and N-epitope-specific probes showed distinct patterns that allowed correlation of the genotypes with the blood group phenotypes. The correlation using S- and s-specific probes was less definite owing to cross-hybridization. An Mspl restriction fragment length polymorphism (RFLP) residing in the E gene was detected in the black population. This RFLP is also carried by all individuals tested who exhibit the S(-)s(-)U- and S(-)s(-)U+ blood groups phenotypes, thereby serving as a useful marker for the S-s- alleles. The site of cleavage resulting in this RFLP was localized to the second intron of the E gene, and cleavage could occur through differential methylation of its two alleles.

A low-incidence red cell antigen JAL associated with two unusual Rh gene complexes.

A multilaboratory investigation during several years has identified a low incidence antigen JAL on the red cells of 7 propositi. JAL appears to be associated with two unusual Rh complexes, one of which produces a depressed C antigen and the other a depressed c antigen. Family studies strongly suggest that the JAL antigen is encoded by the RH locus. Anti-JAL has been implicated in haemolytic disease of the newborn and is thus considered to be a clinically significant antibody.

Kg, a new low-frequency red cell antigen responsible for hemolytic disease of the newborn.

Hemolytic disease of the newborn in a Japanese infant led to studies which indicate that an antibody detected in the maternal serum is recognizing a hitherto unknown red cell antigen. The antigen, which we have named Kg, was found in two generations of the family and it is inherited as a Mendelian dominant character. The maternal serum failed to react with the red cells of more than 600 random Japanese blood donors, 64 red cell samples known to possess low-frequency antigens and all but one of 75 red cell samples known to lack high-frequency antigens without recognized low-frequency antigens. The father's red cells were tested extensively for established low-frequency antigens; only Dia was demonstrated and the maternal antibody was shown not to contain anti-Dia.

Emm. A red cell antigen of very high frequency.

Anti-Emm, an antibody to a public red cell antigen, was found in the serums of four Emm-propositi and in the serum of the Emm-brother of one of them. Three of the propositi were men, and their antibodies, as well as that of the compatible brother, were believed to be "naturally occurring." Since there are two Emm-siblings in one family, Emm is probably an inherited characteristic.

Adaptation to chronic hyperoxia. Biochemical effects and the response to subsequent lethal hyperoxia.

Adult rats were exposed to room air, 50%, 65%, or 80% oxygen for 6 wk. Subsets were sacrificed periodically in order to establish alterations in growth parameters and lung antioxidant responses. Prolonged exposure to 50% or 65% oxygen did not result in weight loss or changes in lung-to-body weight ratios relative to control values. Treatment with 50% oxygen produced delayed increases in nonprotein sulfhydryl (NPSH) content and catalase (CAT) activity, while treatment with 65% oxygen produced delayed increases in NPSH, CAT, and glutathione peroxidase (GPx) content. Rats treated for 6 wk with either 50% or 65% oxygen died significantly earlier than room-air control animals when these groups were subsequently exposed to 100% oxygen. Rats exposed to 80% oxygen had significantly decreased body weight, increased lung-to-body weight ratios, and increased levels of NPSH, CAT, GPx, total superoxide dismutase, and glutathione reductase by 11 days of treatment. At 6 wk they had significantly altered growth parameters and increased GPx catalase, and NPSH levels. Their final antioxidant profile was not significantly different from 65% oxygen-exposed rats. However, these animals survived significantly longer than any group when exposed to 100% oxygen. In summary, lower concentrations of sublethal hyperoxia (less than or equal to 65%) were capable of eliciting significant antioxidant enzyme responses. Levels of antioxidant enzymes in the lungs of rats chronically exposed to sublethal hyperoxia did not appear to be solely responsible for enhanced survival in subsequent lethal hyperoxia.

Membrane glycophorins in Sta blood group erythrocytes.

Structural and immunochemical studies of glycophorins isolated from erythrocytes of an individual homozygous for the M Sta blood group phenotype are described. Reactivities with specific monoclonal antibodies indicated that two major M and N glycophorins were present. The M and N Sta glycophorins were resolved by Lens culinaris lectin affinity chromatography. The N species was not held on the lectin but the M species, like control alpha glycophorins, was retained and could be eluted with alpha-methylmannoside. The two proteins were present in almost equimolar amounts. Studies of the CNBr fragments provided evidence that the structure of M Sta glycophorin is the same as that of the usual M alpha glycophorin but that the N Sta glycophorin is a variant. The amino-terminal octapeptides of the M and N species were similar in amino acid and carbohydrate composition to those isolated, respectively, from M and N alpha glycophorins. The studies focused on CNBr glycopeptide B that, in control alpha glycophorins, extends from amino acid residues 9 to 81. The fragment from the M species exhibited properties identical to those of the corresponding fragment of control alpha glycophorins in terms of size, chromatographic behavior, amino acid and carbohydrate contents and compositions, the presence of O-glycosidically linked saccharides and a single Asn-linked carbohydrate unit. The structures of the O-linked units were inferred experimentally to be NeuAc(alpha 2,3)Gal-(beta 1,3)GalNAc and NeuAc(alpha 2,3)Gal(beta 1,3) [NeuAc(alpha 2,6)]GalNAc, present in a ratio similar to that found in controls; and the Asn-linked unit also appeared to be as in the control. The tryptic glycopeptide pattern of the M Sta glycophorin CNBr fragment B was identical to the pattern of the corresponding control fragment, and the composition of the tryptic peptides suggested sequence identity with the control fragment. In contrast, the N Sta glycophorin yielded two CNBr glycopeptides B; both contained fewer amino acid residues and virtually lacked Man and GlcNAc, indicating the absence of the Asn-linked carbohydrate. The much decreased levels of these carbohydrates in the intact N protein, corroborated the latter finding. The O-glycosidic saccharides appeared similar to those found in control alpha glycophorins. However, the tryptic glycopeptide pattern of the variant differed from control M or N alpha glycophorins, suggesting a deletion of a large segment of the molecule near residues 40-61 and/or a substitution of methionine for a residue upstream from residue 40.(ABSTRACT TRUNCATED AT 400 WORDS)