A novel pathway of levodopa metabolism by commensal Bifidobacteria.

The gold-standard treatment for Parkinson's disease is levodopa (L-DOPA), which is taken orally and absorbed intestinally. L-DOPA must reach the brain intact to exert its clinical effect; peripheral metabolism by host and microbial enzymes is a clinical management issue. The gut microbiota is altered in PD, with one consistent and unexplained observation being an increase in Bifidobacterium abundance among patients. Recently, certain Bifidobacterium species were shown to have the ability to metabolize L-tyrosine, an L-DOPA structural analog. Using both clinical cohort data and in vitro experimentation, we investigated the potential for commensal Bifidobacteria to metabolize this drug. In PD patients, Bifidobacterium abundance was positively correlated with L-DOPA dose and negatively with serum tyrosine concentration. In vitro experiments revealed that certain species, including B. bifidum, B. breve, and B. longum, were able to metabolize this drug via deamination followed by reduction to the compound 3,4-dihydroxyphenyl lactic acid (DHPLA) using existing tyrosine-metabolising genes. DHPLA appears to be a waste product generated during regeneration of NAD +. This metabolism occurs at low levels in rich medium, but is significantly upregulated in nutrient-limited minimal medium. Discovery of this novel metabolism of L-DOPA to DHPLA by a common commensal may help inform medication management in PD.

Cross-feeding between intestinal pathobionts promotes their overgrowth during undernutrition.

Child undernutrition is a global health issue associated with a high burden of infectious disease. Undernourished children display an overabundance of intestinal pathogens and pathobionts, and these bacteria induce enteric dysfunction in undernourished mice; however, the cause of their overgrowth remains poorly defined. Here, we show that disease-inducing human isolates of Enterobacteriaceae and Bacteroidales spp. are capable of multi-species symbiotic cross-feeding, resulting in synergistic growth of a mixed community in vitro. Growth synergy occurs uniquely under malnourished conditions limited in protein and iron: in this context, Bacteroidales spp. liberate diet- and mucin-derived sugars and Enterobacteriaceae spp. enhance the bioavailability of iron. Analysis of human microbiota datasets reveals that Bacteroidaceae and Enterobacteriaceae are strongly correlated in undernourished children, but not in adequately nourished children, consistent with a diet-dependent growth synergy in the human gut. Together these data suggest that dietary cross-feeding fuels the overgrowth of pathobionts in undernutrition.

Dietary Intervention Reverses Fatty Liver and Altered Gut Microbiota during Early-Life Undernutrition.

Nonalcoholic fatty liver disease (NAFLD), largely studied as a condition of overnutrition, also presents in undernourished populations. Like NAFLD, undernutrition disrupts systemic metabolism and has been linked to gut microbiota dysbiosis. Indeed, chronic exposures to fecal microbes contribute to undernutrition pathology in regions with poor sanitation. Despite a growing prevalence of fatty liver disease, the influence of undernutrition and the gut microbiota remain largely unexplored. Here, we utilize an established murine model (C57BL/6J mice placed on a malnourished diet that received iterative Escherichia coli/Bacteroidales gavage [MBG mice]) that combines a protein/fat-deficient diet and iterative exposure to specific, fecal microbes. Fecal-oral contamination exacerbates triglyceride accumulation in undernourished mice. MBG livers exhibit diffuse lipidosis accompanied by striking shifts in fatty acid, glycerophospholipid, and retinol metabolism. Multiomic analyses revealed metabolomic pathways linked to the undernourished gut microbiome and hepatic steatosis, including phenylacetate metabolism. Intriguingly, fatty liver features were observed only in the early-life, but not adult, MBG model despite similar liver metabolomic profiles. Importantly, we demonstrate that dietary intervention largely mitigates aberrant metabolomic and microbiome features in MBG mice. These findings indicate a crucial window in early-life development that, when disrupted by nutritional deficiency, may significantly influence liver function. Our work provides a multifaceted study of how diet and gut microbes inform fatty liver progression and reversal during undernutrition.IMPORTANCE Nonalcoholic fatty liver disease (NAFLD) remains a global epidemic, but it is often studied in the context of obesity and aging. Nutritional deficits, however, also trigger hepatic steatosis, influencing health trajectories in undernourished pediatric populations. Here, we report that exposure to specific gut microbes impacts fatty liver pathology in mice fed a protein/fat-deficient diet. We utilize a multiomics approach to (i) characterize NAFLD in the context of early undernutrition and (ii) examine the impact of diet and gut microbes in the pathology and reversal of hepatic steatosis. We provide compelling evidence that an early-life, critical development window facilitates undernutrition-induced fatty liver pathology. Moreover, we demonstrate that sustained dietary intervention largely reverses fatty liver features and microbiome shifts observed during early-life malnutrition.

Two-year follow-up of patients with septic shock presenting with low HDL: the effect upon acute kidney injury, death and estimated glomerular filtration rate.

BACKGROUND: Sepsis is associated with decreased levels of high-density lipoprotein (HDL) cholesterol. HDL has anti-inflammatory properties, and the use of Apo A-I mimetic peptides is associated with renal function improvement in animal models of sepsis. However, it is not known whether decreased HDL level results in impaired renal function in human sepsis. We investigated whether low levels of HDL conferred an increased risk of sepsis-associated acute kidney injury (AKI) or long-term decreased estimated glomerular filtration rate (eGFR) after sepsis. METHODS: HDL concentration (mg dL-1 ) was measured in plasma samples from 180 patients with septic shock at admission to the Emergency Department (ED). We divided the patients using median HDL as a cut-off value and assessed the frequency of sepsis-associated AKI and long-term decreased eGFR after sepsis. Univariate and multivariate analyses were performed. RESULTS: Patients with low HDL had a significantly greater frequency of KDIGO 2 or 3 sepsis-associated AKI [39/90 (43.3%) vs. 12/90 (13.3%), P < 0.001] and decreased long-term eGFR [24/58 (41.4%) vs. 11/57 (19.3%), P = 0.018] compared to those with high HDL. The adjusted OR for sepsis-associated AKI and decreased eGFR after sepsis in the lower HDL group was 2.80 (95% CI 1.08-7.25, P = 0.033) and 5.45 (95% CI 1.57-18.93, P = 0.008), respectively. CONCLUSION: Low HDL levels during sepsis are associated with increased risk of sepsis-associated AKI, and/or subsequent decreased eGFR. These results suggest that HDL may be involved and/or may be a marker of kidney injury during and after sepsis.

Improvement of arm movement patterns and endpoint control depends on type of feedback during practice in stroke survivors.

BACKGROUND: A major challenge in stroke rehabilitation is restoration of arm motor function. Therapy-induced improvements in arm function may occur via restoration of premorbid movement patterns (recovery) or development of compensatory movement strategies. However, it is unclear whether the learning benefits of practice might be enhanced by incorporating different forms of feedback, focusing on movement outcomes or on specific arm movement patterns. OBJECTIVE: To determine if manipulation of attentional focus by providing either knowledge of results (KR) feedback, focusing on movement outcomes, or knowledge of performance (KP) feedback, focusing on arm movement patterns during repetitive practice of a pointing movement, may lead to arm motor recovery. METHODS: Twenty-eight chronic stroke survivors were randomly assigned to 2 groups that practiced 10 sessions of 75 pointing movements. During practice, groups received either 20% KR about movement precision or faded (26.6% average) KP about arm joint movements. A nondisabled control group (n = 5) practiced the same task with KR. RESULTS: Motor patterns recovered only in KP, as evidenced by immediate and long-term increases in joint range, better interjoint coordination in early movement phases, and generalization of gains. Improvements in clinical impairment and function were related to decreases in compensation (trunk rotation) and recovery of interjoint coordination in mid-movement phases. CONCLUSIONS: In stroke survivors, when the learners' attention was directed to the movements themselves (KP), motor improvements reflect recovery compared to when attention was directed toward movement outcomes (KR).

Arm reaching improvements with short-term practice depend on the severity of the motor deficit in stroke.

The effects of short-term, constant practice on the kinematics of a multi-joint pointing movement were studied in the hemiparetic arm of 20 chronic patients with unilateral left cerebro-vascular accident (CVA) and in 10 age- and sex-matched healthy individuals. Practice consisted of a single session of 70 pointing movements made with the right arm. Movements were made from a target located beside the body to one in the contralateral workspace, in front of the body. Vision of the final hand position was allowed after every 5th trial. At the beginning of practice, stroke patients made slower, less precise and more segmented movements, characterised by smaller active ranges of elbow and shoulder motion, disrupted elbow-shoulder coordination, as well as greater trunk movement compared with healthy subjects. With practice, healthy subjects and some patients made faster and more precise movements. These tendencies were revealed only after many repetitions (up to 55 for those with severe hemiparesis), whereas changes in healthy individuals occurred after fewer trials (approximately 20). In addition, the patients decreased movement segmentation with practice. In healthy subjects, faster movement times may be attributed to better shoulder/elbow movement timing in the first half of the reach, whereas improvement of precision was not correlated with any changes in the movement variables. In patients, improvements were accomplished differently depending on arm motor severity. For some patients with mild-to-moderate clinical symptoms, practice resulted in better timing of shoulder/elbow movements with less trunk rotation in middle to late reach. Patients with more severe impairment also improved shoulder/elbow movement timing in mid-reach but used more compensatory trunk rotation. The results suggest that even one session of repetitive practice of a multi-joint pointing task leads to improvements in movement performance-based outcome measures, but the mechanisms of improvement may vary with the individual's level of motor impairment.

Interjoint coordination dynamics during reaching in stroke.

A technique is described that characterizes the dynamics of the interjoint coordination of arm reaching movements in healthy subjects (n=10) and in patients who had sustained a left-sided cerebrovascular accident (n=18). All participants were right-handed. Data from the affected right arm of patients with stroke were compared with those from the right arm of healthy subjects. Seated subjects made 25 pointing movements in a single session. Movements were made from an initial target located ipsilaterally to the right arm beside the body, to a final target located in front of the subject in the contralateral arm workspace. Kinematic data from the finger, wrist, elbow, both shoulders and sternum were recorded in three dimensions at 200 Hz with an optical tracking system. Analysis of interjoint coordination was based on the patterns of temporal delay between rotations at two adjacent joints (shoulder and elbow). The data were reduced to a single graph (Temporal Coordination or TC index) integrating the essential temporal characteristics of joint movement (the angular displacements, velocities and timing). TC segments, duration and amplitude, were analysed. The analysis was sensitive to the differences in interjoint coordination between healthy subjects and patients with arm motor deficits. In patients, the temporal coordination between elbow and shoulder movements was disrupted from the middle to the end of the reach. More specifically, in mid-reach, all patients had difficulty coordinating elbow flexion with shoulder horizontal adduction. In addition, patients with severe arm hemiparesis had difficulty changing elbow movement direction from flexion to extension and in coordinating this change with shoulder movement. At the end of the reach, patients with severe hemiparesis had deficits in the execution of elbow extension while all patients had impaired coordination of elbow extension and shoulder horizontal adduction. In addition, active ranges of joint motions were significantly decreased in the stroke compared to the healthy subjects. Finally, TC analysis revealed significant relationships between specific aspects of disrupted interjoint coordination and the level of motor impairment, suggesting that it may be a useful tool in the identification of specific movement coordination deficits in neurological impaired populations that can be targeted in treatment for arm motor recovery.

Compensatory strategies for reaching in stroke.

A major prerequisite for successful rehabilitation therapy after stroke is the understanding of the mechanisms underlying motor deficits common to these patients. Studies have shown that in stroke patients multijoint pointing movements are characterized by decreased movement speed and increased movement variability, by increased movement segmentation and by spatial and temporal incoordination between adjacent arm joints with respect to healthy subjects. We studied how the damaged nervous system recovers or compensates for deficits in reaching, and correlated reaching deficits with the level of functional impairment. Nine right-hemiparetic subjects and nine healthy subjects participated. All subjects were right-hand dominant. Data from the affected arm of hemiparetic subjects were compared with those from the arm in healthy subjects. Seated subjects made 40 pointing movements with the right arm in a single session. Movements were made from an initial target, for which the arm was positioned alongside the trunk. Then the subject lifted the arm and pointed to the final target, located in front of the subject in the contralateral workspace. Kinematic data from the arm and trunk were recorded with a three-dimensional analysis system. Arm movements in stroke subjects were longer, more segmented, more variable and had larger movement errors. Elbow-shoulder coordination was disrupted and the range of active joint motion was decreased significantly compared with healthy subjects. Some aspects of motor performance (duration, segmentation, accuracy and coordination) were significantly correlated with the level of motor impairment. Despite the fact that stroke subjects encountered all these deficits, even subjects with the most severe motor impairment were able to transport the end-point to the target. All but one subject involved the trunk to accomplish this motor task. In others words, they recruited new degrees of freedom typically not used by healthy subjects. The use of compensatory strategies may be related to the degree of motor impairment: severely to moderately impaired subjects recruited new degrees of freedom to compensate for motor deficits while mildly impaired subjects tended to employ healthy movement patterns. We discuss the possibility that there is a critical level of recovery at which patients switch from a strategy employing new degrees of freedom to one in which motor recovery is produced by improving the management of degrees of freedom characteristic of healthy performance. Our data also suggest that stroke subjects may be able to exploit effectively the redundancy of the motor system.

The diagnostic value of intradermal tests with penicilloyl-dextran and aspiryl-polylysine.

Intradermal tests (i.d.t.) with penicillin (P) and penicilloyl-dextran (P-Dex) were performed in 32 penicillin allergic patients and in 37 controls with allergy to other drugs. The incidence of the positive i.d.t. was much higher in the group of penicillin allergy of immediate type (17 of 18 patients) than in the group of penicillin allergy of half-late or late type (4 of 14 cases). I.d.t. with P and P-Dex were concordant in 75% of cases. Among 28 patients with penicillin allergy tested with both reagents, i.d.t. positive only to P-Dex were found in two cases. The utilisation of i.d.t. with P-Dex or penicilloyl-polylysine (PPL) in penicillin allergy furnishes, therefore, only very modest additional information besides that furnished by i.d.t. performed with plain penicillin. I.d.t. with aspiryl-polylysine (APL) were performed in 23 patients with aspirin allergy/intolerance and in 60 controls with allergy to other drugs. 70% of the individuals with aspirin allergy and 27% of the patients with allergy to other drugs showed positive i.d.t. to APL. Although the specificity of i.d.t. with APL in aspirin allergy seems lower than that of i.d.t. with P-Dex and PPL in penicillin allergy, a positive cutaneous test with APL may constitute an important argument for the allergic nature of an aspirin adverse reaction evoking an allergic mechanism.

On the relation between the formation of antihapten and anti-carrier protein antibodies in the immune response induced by conjugates of various haptens with homologous serum proteins.

Rabbits inoculated with conjugates of homologous serum with ortho-aminophenol (OAP), para-aminophenol (PAP), 3-hydroxy-sulfanilamide (3-HS), DOPA and alpha-methyldopa (alpha MD), 5-aminosalicylic acid (5-ASA) and 3-hydroxy-para-aminobenzoic acid (3-HPABA) give rise to both hapten specific and anti-carrier protein antibodies. By contrast, the conjugates of homologous serum with penicilloyl (P) hapten induced only antihapten antibodies. The conclusion is, therefore, drawn that the formation of anti-carrier antibodies upon inoculation with conjugates of various haptens appears to be dependent on the nature of hapten in the conjugate. The discrepancy between the behaviour of P conjugates, on the one hand, and the DOPA and alpha MD conjugates, on the other hand, is in agreement with the observation from the clinical immunology, which has signaled a high incidence of antibodies with autoantibody character in patients taking DOPA and alpha MD, but rarely, if ever, such antibodies have been found in patients treated with penicillin.

Haptenic properties of drug metabolites with ortho- or para-aminophenolic structure.

3-Hydroxy-sulfanilamide (3-HS), a biotransformation product of sulfanilamide, 3-hydroxy-para-aminobenzoic acid (3- HPABA ), a biotransformation product of procaine and para-aminobenzoic acid, as well as 5-aminosalicylic acid (5-ASA), a bacterial split product of salicylazosulfapyridine (Sulfasalazine, Salazopyrin ) in the gut, were shown to react spontaneously with and to bind covalently to proteins by simple incubation at alkaline pH. When administered incorporated in incomplete Freund adjuvant into rabbits, the conjugates of these haptens with normal rabbit serum proteins (NRS) give rise to both hapten specific and anti-NRS antibodies. The haptenic properties of these drug metabolites are apparently due to the presence in their molecules of an ortho- or para- aminophenolic structure that renders them capable to generate spontaneously highly reactive quinoneimines . Extensive cross immunological reactions were found between 3-HS, 3- HPABA and ortho-aminophenol (OAP) immunological systems, all of these haptens belonging to the ortho- aminophenolic (ortho- quinoneiminic ) system. Cross reactions were also found between 5-ASA and para-aminophenol (PAP) immunological systems, both of these haptens belonging to the para- aminophenolic (para- quinoneiminic ) system. No cross reactions were, however, found between ortho- and para-aminophenol immunological systems. The so-called "para" group of aminoaromatic compounds is not therefore immunochemically homogeneous. Some compounds included usually in this group give rise in the body to haptens belonging to the ortho- aminophenolic systems, while others belong really to the para- aminophenolic system. Antibodies against NRS proteins which served as carrier in the conjugates were formed by most animals inoculated with such conjugates. (ABSTRACT TRUNCATED AT 250 WORDS)

Spontaneous formation of immunogenic conjugates by 3-hydroxy-para-aminobenzoic acid--a biotransformation product of procaine and para-aminobenzoic acid.

3-Hydroxy-para-aminobenzoic acid (3-HPABA), a biotransformation product of procaine and para-aminobenzoic acid, is capable of binding covalently to proteins by simple incubation at alkaline pH. When administered incorporated in Freund adjuvant to rabbits, 3-HPABA conjugates with normal rabbit serum proteins (3-HPABA-NRS) give rise to both hapten specific antibodies and anti-NRS antibodies. Cross immunological reactions were found between 3-HPABA and ortho-aminophenol (OAP) and 3-hydroxy-sulfanilamide (3-HS) immunological systems, but not between 3-HPABA and para-aminophenol (PAP) and 5-aminosalicyclic acid (5-ASA) immunological systems. The significance of these findings for the explanation of procaine allergy and the possible use of 3-HPABA conjugates for skin testing in procaine hypersensitivity is discussed.

Spontaneous formation of immunogenic conjugates by 5-aminosalicylic acid--a biotransformation product of salicylazosulfapyridine.

5-Aminosalicylic acid (5-ASA) reacts readily with proteins at alkaline pH and forms conjugates capable to induce hapten specific antibodies in rabbits. Extensive immunological cross reactions have been found between 5-ASA and para-aminophenol (PAP) immunological systems, but no cross reactions were found between 5-ASA and orto-aminophenol (OAP) immunological systems. Moderate cross reactions were found, occasionally between 5-ASA, 3-hydroxy-sulfanilamide (3-HS) and alpha-methyldopa (alpha MD) immunological systems. Antibodies directed against normal rabbit serum (NRS) proteins, which served as carrier in the 5-ASA conjugates used for immunization, were found in the sera of three out of the six rabbits inoculated with such conjugates. Antibodies directed against heterologous proteins were found in all of these sera, including one serum which lacked hapten specific antibodies and the three sera lacking antibodies directed against NRS proteins. The significance of these findings for the interpretation of the immuno-allergic reactions induced by sulfasalazine is discussed.

Spontaneous formation of conjugates of alpha-methyl-DOPA and DOPA with serum proteins and experimental production of specific antibodies against these haptens.

Alpha-methyldopa (alpha MD) and levodopa (DOPA) react readily with serum proteins at alkaline pH and form stable conjugates with them. Hapten specific antibodies in high titers are produced in rabbits by inoculation of the conjugates formed by alpha MD and DOPA with normal rabbit serum (alpha MD-NRS and DOPA-NRS), incorporated in incomplete Fruend adjuvant. No hapten specific antibodies were raised in rabbits inoculated with alpha MD as such, incorporated in adjuvant, with alpha MD mixed with autologous serum 24 hrs before inoculation, or with alpha MD conjugates with autologous serum administered without adjuvant. Hapten specific antibodies were, however, found in one of six rabbits inoculated with alpha MD conjugates with homologous serum proteins given without adjuvant. Extensive crossreactivity was found between alpha MD and DOPA immunological systems. High titers of antibodies directed against normal rabbit serum proteins as well as against human gamma globulin were also found in the sera of the rabbits immunized with alpha MD-NRS and DOPA-NRS conjugates incorporated in Freund adjuvant. A close correlation was found between the titers of hapten specific antibodies and of anti-protein antibodies.

Penicilloyl-dextran conjugates as reagents for skin testing in penicillin allergy.

A procedure for implanting amino groups in the dextran molecule is described. The aminated polymer prepared from a dextran sample with a molecular weight of 20,000 was found to contain approximately 16 amino groups per molecule. Similarly to polylysine, the aminated dextran may be penicilloylated by incubation with penicillin at high pH. The penicilloyl-dextran (P-Dex) conjugate prepared in this way was found to contain approximately 8 penicilloyl (PO) groups per molecule. P-Dex was found practically nonimmunogenic in rabbits and showed itself antigenically active in precipitin, passive hemagglutination and passive cutaneous anaphylaxis (PCA) tests performed with a PO specific antiserum. A good correlation was found between the skin tests performed parallelly with P-Dex and penicilloyl-polylysine (PPL) in four subjects with penicillin allergy.

A comparative study of the immunogenicity of some penicillins and of their cross reactions.

The immunogenicity of four penicillin preparations was comparatively investigated by titration of penicilloyl (PO) specific hemagglutinating antibodies found in sera of rabbits inoculated with equimolecular amounts of these penicillins incorporated in Freund incomplete adjuvant. Benzylpenicillin (BP) was shown to be endowed with the highest immunogenicity, while oxacillin (OX), ampicillin (AM) and Amoxil (AX) were found significantly less immunogenic. By examining comparatively the titers of hemagglutination reactions given by the sera of each animal with erythrocytes coated with the penicillin used for its immunization as well as with the other penicillins investigated, important cross immunological reactions were found in all of the cases. Interestingly, the cross reactions produced by AM- and AX-specific antisera with heterologous penicillins were apparently higher than the reactions obtained with the homologous penicillin. No significant differences were found concerning the ability of the four penicillins to form PO conjugates with BGG. Thus, the differences in immunogenicity exhibited by the four penicillin preparations may not be explained in this way.