Faculty Bibliography

Introduction: People with schizophrenia have been reported to show deficits in tests of olfactory function. DNA methylation and GABAergic input have been implicated in biochemical processes controlling odor in animal studies, but this has not been investigated in human studies. Methods: In a study of measures of DNA methylation and GABAergic mRNAs in lymphocytes, we also measured odor identification and discrimination with the Sniffin"™ Sticks battery in 58 patients with chronic schizophrenia (CSZ) and 48 controls. mRNAs in lymphocytes were assessed by qPCR using TaqManTM probes. Cognition was assessed by the MATRICS battery (Measurement and Treatment Research to Improve Cognition in Schizophrenia) in CSZ and controls, and symptoms in CSZ were assessed by PANSS scale (Positive and Negative Symptom Scale). The relationships of odor deficits with mRNA, cognition, and symptoms were explored by correlation analysis. Variables which significantly differentiated CSZ from controls were explored by logistic regression. Results: Overall, CSZ showed significantly (P≤.001) lower scores on odor discrimination compared to controls, with a moderate effect size, but no difference in odor identification. Deficits in odor discrimination, which has not been standardly assessed in many prior studies, strongly differentiated CSZ from controls. In logistic regression analysis, odor discrimination, but not odor identification, was a significant variable predicting schizophrenia versus control class membership. This is the first study to report relationship between odor deficits and DNA methylation and GABAergic mRNAs in blood cells of human subjects. There were negative correlations of odor identification with DNA methylation enzymes mRNAs and significant negative correlations with odor discrimination and GABAergic mRNAs. Lower odor scores were significantly associated with lower cognitive scores on the MATRICS battery in CSZ but not control subjects. In CSZ, lower odor scores were significantly associated with negative symptom scores, while higher odor identification scores were associated with PANNS Excitement factor. Discussion: Odor discrimination was a more powerful variable than odor identification in discriminating CSZ from controls and should be used more regularly as an odor measure in studies of schizophrenia. The substantive meaning of the negative correlations of odor discrimination and GABAergic mRNA variables in peripheral lymphocytes of CSZ needs more investigation and comparison with results in neural tissue.

Background: Patients with schizophrenic have been reported to show deficits in various measures odor perception but odor discrimination has not been standardly assessed. DNA methylation and GABAergic input have been implicated in biochemical process controlling odor in animal studies, but this has not been investigated in human studies. Some studies have related cognitive deficits in schizophrenia to odor deficits but none have used the MATRICS battery to investigate this question.
Method(s): In a study of DNA methylation and GABAergic mRNAs in lymphocytes we also measured odor identification and discrimination with the Sniff and Sticks battery in 58 patients with chronic schizophrenia (CSZ) and 48 non-psychiatric controls (NPC). mRNAs in lymphocytes were assessed by qPCR using TaQMan probes. Cognition was assessed by the MATRICS battery in CSZ and NPC and symptoms in CSZ were assessed by PANSS scale. The relationship of odor deficits to mRNA levels and MATRICS scores and symptoms was explored by correlation analysis.
Result(s): CSZ showed significant deficits compared to NPC in odor identification (P = 0.011, Cohen's d = 0.50)), but much larger deficits in discrimination (P < 0.001, d = 1.01). In step down regression analysis odor discrimination but not odor identification had significant beta weight for classify patients into the CSZ vs NPC group. There were significant negative correlations (r = -33 to -.68) of odor identification with DNMT1 mRNAs, and significant negative correlations with odor discrimination and GABAergic mRNAs in CSZ subjects (-.38 to -.42). Odor discrimination scores correlated significantly (P = 0.02 to P = 0.009) with several Matrics Domain scores in CSZ subjects but not NPC; there was a sex effect and these correlations were stronger in female than male CSZ.
Conclusion(s): Odor discrimination deficits, which has not been consistently evaluated in schizophrenia studies, showed the strongest differentiation between patients with schizophrenia and controls. This is the first study to report relationship between odor deficits and DNMT and GABAergic mRNAs in human subjects. However, the negative correlations of odor scores with lymphocyte mRNA levels may not necessarily reflect neuronal processes

Background/UNASSIGNED:Cannabis use is a component risk factor for the manifestation of schizophrenia. The biological effects of cannabis include effects on epigenetic systems, immunological parameters, in addition to changes in cannabinoid receptors 1 and 2, that may be associated with this risk. However, there has been limited study of the effects of smoked cannabis on these biological effects in human peripheral blood cells. We analyzed the effects of two concentrations of tetrahydrocannabinol (THC) vs. placebo in lymphocytes of a subset of participants who enrolled in a double-blind study of the effects of cannabis on driving performance (outcome not the focus of this study). Methods/UNASSIGNED:-tests, or non-parametric equivalents for those values which were not normally distributed. Results/UNASSIGNED:= 0.056). The higher 13.4% THC group had significantly increased CB2 mRNA levels than the 5.9% concentration group at several post drug administration time points and showed trends for difference in effects for between 5.9 and 13.4% THC groups for other mRNAs. TET3 mRNA levels were higher in the 13.4% THC group at 55 min post-cannabis ingestion. When the high and lower concentration THC groups were combined, none of the differences in mRNA levels from placebo remained statistically significant. Changes in THC blood levels were not related to changes in mRNA levels. Conclusion/UNASSIGNED:Over the time course of this study, CB2 mRNA increased in blood lymphocytes in the high concentration THC group but were not accompanied by changes in immunological markers. The changes in DNMT and TET mRNAs suggest potential epigenetic effects of THC in human lymphocytes. Increases in DNMT methylating enzymes have been linked to some of the pathophysiological processes in schizophrenia and, therefore, should be further explored in a larger sample population, as one of the potential mechanisms linking cannabis use as a trigger for schizophrenia in vulnerable individuals. Since the two THC groups did not differ in post-smoking blood THC concentrations, the relationship between lymphocytic changes and the THC content of the cigarettes remains to be determined.

Some of the biochemical abnormalities underlying schizophrenia, involve differences in methylation and methylating enzymes, as well as other related target genes. We present results of a study of differences in mRNA expression in peripheral blood lymphocytes (PBLs) and post-mortem brains of chronic schizophrenics (CSZ) and non-psychotic controls (NPC), emphasizing the differential effects of sex and antipsychotic drug treatment on mRNA findings. We studied mRNA expression in lymphocytes of 61 CSZ and 49 NPC subjects using qPCR assays with TaqMan probes to assess levels of DNMT, TET, GABAergic, NR3C1, BDNF mRNAs, and several additional targets identified in a recent RNA sequence analysis. In parallel we studied DNMT1 and GAD67 in samples of brain tissues from 19 CSZ, 26 NPC. In PBLs DNMT1 and DNMT3A mRNA levels were significantly higher in male CSZ vs NPC. No significant differences were detected in females. The GAD1, NR3C1 and CNTNAP2 mRNA levels were significantly higher in CSZ than NPC. In CSZ patients treated with clozapine, GAD-1 related, CNTNAP2, and IMPA2 mRNAs were significantly higher than in CSZ subjects not treated with clozapine. Differences between CSZ vs NPC in these mRNAs was primarily attributable to the clozapine treatment. In the brain samples, DNMT1 was significantly higher and GAD67 was significantly lower in CSZ than in NPC, but there were no significant sex differences in diagnostic effects. These findings highlight the importance of considering sex and drug treatment effects in assessing the substantive significance of differences in mRNAs between CSZ and NPC.

Homochirality of DNA and prevalent chirality of free and protein-bound amino acids in a living organism represents the challenge for modern biochemistry and neuroscience. The idea of an association between age-related disease, neurodegeneration, and racemization originated from the studies of fossils and cataract disease. Under the pressure of new results, this concept has a broader significance linking protein folding, aggregation, and disfunction to an organism's cognitive and behavioral functions. The integrity of cognitive function is provided by a delicate balance between the evolutionarily imposed molecular homo-chirality and the epigenetic/developmental impact of spontaneous and enzymatic racemization. The chirality of amino acids is the crucial player in the modulation the structure and function of proteins, lipids, and DNA. The collapse of homochirality by racemization is the result of the conformational phase transition. The racemization of protein-bound amino acids (spontaneous and enzymatic) occurs through thermal activation over the energy barrier or by the tunnel transfer effect under the energy barrier. The phase transition is achieved through the intermediate state, where the chirality of alpha carbon vanished. From a thermodynamic consideration, the system in the homo-chiral (single enantiomeric) state is characterized by a decreased level of entropy. The oscillating protein chirality is suggesting its distinct significance in the neurotransmission and flow of perceptual information, adaptive associative learning, and cognitive laterality. The common pathological hallmarks of neurodegenerative disorders include protein misfolding, aging, and the deposition of protease-resistant protein aggregates. Each of the landmarks is influenced by racemization. The brain region, cell type, and age-dependent racemization critically influence the functions of many intracellular, membrane-bound, and extracellular proteins including amyloid precursor protein (APP), TAU, PrP, Huntingtin, α-synuclein, myelin basic protein (MBP), and collagen. The amyloid cascade hypothesis in Alzheimer's disease (AD) coexists with the failure of amyloid beta (Aβ) targeting drug therapy. According to our view, racemization should be considered as a critical factor of protein conformation with the potential for inducing order, disorder, misfolding, aggregation, toxicity, and malfunctions.

Background: There is evidence from animal and in-vitro studies that THC can affect cannabinoid receptors (1 and2) in brain and periphery and influence immunological makers, and limited evidence from animal studies that it may affect epigenetic related methylation processes. THC ingestion has also been reported as a trigger for inducing schizophrenia in venerable individuals. This is less direct work on these effects in human cannabis smokers investigating these types of chemical biomarkers in peripheral blood cells. The present study further evaluated whether smoked cannabis in human subjects produced changes in cannabinoid receptors, biomarkers for DNA methylation cycle and immunorelated gene mRNA expression.
Method(s): 23 Subjects participated in an experiment in which they smoked cannabis cigarettes with one of two doses of marijuana (5.3% or 13.4% THC) or placebo (0.02%) and were evaluated driving abilities. Blood samples were drawn at baseline and several times after smoking. Plasma and WBC (PMCs) were separated and stored at -80degreeC until further analysis. Samples were analyzed for mRNA content for cannabinoid receptors 1(CBR1) and 2(CBR2), methylation and demethylating enzymes (DNMT, TET), glucocorticoid receptor (NRC3) and immunological markers (IL1B, TNFalpha) by qPCR using TaqMan probes. The results were correlated with THC whole blood levels and TCOOH baseline levels. Statistical analyzed used analysis of variance and covariance and t-test, or non-parametric equivalents for those values which were not normally distributed.
Result(s): There were no difference in background baseline characteristics of the subjects except that the higher dose THC group was older than the low dose and placebo groups, and the low dose THC group had higher baseline CBR2 mRNA levels. Both the 5.9 and 13.4 THC groups showed increased THC levels over the next 2 hours and then decreased toward baseline; the 13.4 THC dose still showed a higher THC levels than placebo at four hours. However, there were no significant differences between THC levels between the 5.9 and 13.4 doses at any time point. At the 4-hour time point after drug administration the 13.4% THC group had higher CBR2 (P = .021) and DNMT3A (P = .027) mRNA levels than the placebo group and DNMT1 mRNA levels showed a trend in the same direction (P = .056). The higher13.4 THC group had significantly higher CBR2 mRNA levels than the 5.9 dose group at several post drug administration time points, and showed trends for difference in effects for between 5.9 and 13.4 THC groups for other mRNAs. TET3 mRNA levels were higher in the 13.4 THC group at 55 minutes post-drug ingestion. When the high and lower dose THC groups were combined, none of the differences in mRNA levels from placebo remain statistically significant. Changes in THC plasma levels were not related to changes in mRNA levels.
Conclusion(s): Over the time course of this study CBR2 levels increased in human PMCs in the high dose THC group but where not accompanied by changes in immunological markers. The changes in DNMT and TET mRNAs suggest potential epigenetic effects of THC in human PMCs. Increases in DNMT metylating enzymes have been linked to some of the pathophysiological process in schizophrenia and, therefore, should be further explored as one of the potential mechanisms linking cannabism use as a trigger for schizophrenia

Background: Some biochemical abnormalities underlying schizophrenia, involve differences in methylation and methylating enzymes, their target genes, and genes uncovered by RNA seq analysis. We present results of a larger study measuring differences in mRNA in lymphocytes (PBL) and brain of chronic schizophrenics (CSZ) and non-psychotic controls (NPC), examining the differential effect of sex and antipsychotic drug treatment.
Method(s): We studied mRNA in PBL of 61 CSZ and 49 NPC using qPCR assays. We studied DNMT1 and GAD67 in brains of 19 CSZ, 26 NPC, and 19 subjects with bipolar disorder (BP).
Result(s): DNMT1 and DNMT3A mRNAs were significantly (P<.01) higher in PBL only in male CSZ subjects, and females showed a trend in the opposite direction. The GAD1, glucocorticoid receptor (NR3C1) and CNTNPA2 mRNAs were significantly higher in CSZ than NPC. The FPRF3 mRNA was significantly lower in CSZ vs NPC. When clozapine treatment was examined, it was clear that for the GABAergic mRNAs (GAD1, GAD67, GAD25), and for NR3C1-B, CNTNPA2, and IMPA2 mRNAs, that clozapine treated CSZ were primarily responsible for the difference from NPC. DNMT1 was significantly higher and GAD67 significantly lower in frontal cortex of brains of CSZ and BP than NPC but there were no sex different effects in this older age brain sample.
Conclusion(s): It is important to consider sex and antipsychotic drug treatment in comparing mRNA levels in schizophrenic patients to controls, since some of the differences are only present in male subjects and other may be explained by clozapine treatment rather than primarily diagnosis. Supported By: NIH 1R01MH101043, and Philanthropic Grant Keywords: Schizophrenia, mRNA, Sex-Specific, Clozapine
Copyright

Biochirality is the subject of distinct branches of science, including biophysics, biochemistry, the stereochemistry of protein folding, neuroscience, brain functional laterality and bioinformatics. At the protein level, biochirality is closely associated with various post-translational modifications (PTMs) accompanied by the non-equilibrium phase transitions (PhTs ^NE). PTMs ^NE support the dynamic balance of the prevalent chirality of enzymes and their substrates. The stereoselective nature of most biochemical reactions is evident in the enzymatic (Enz) and spontaneous (Sp) PTMs (PTMs ^Enz and PTMs ^Sp) of proteins. Protein chirality, which embraces biophysics and biochemistry, is a subject of this review. In this broad field, we focus attention to the amyloid-beta (Aβ) peptide, known for its essential cellular functions and associations with neuropathology. The widely discussed amyloid cascade hypothesis (ACH) of Alzheimer's disease (AD) states that disease pathogenesis is initiated by the oligomerization and subsequent aggregation of the Aβ peptide into plaques. The racemization-induced aggregation of protein and RNA have been extensively studied in the search for the contribution of spontaneous stochastic stereo-specific mechanisms that are common for both kinds of biomolecules. The failure of numerous Aβ drug-targeting therapies requires the reconsolidation of the ACH with the concept of PTMs ^Sp. The progress in methods of chiral discrimination can help overcome previous limitations in the understanding of AD pathogenesis. The primary target of attention becomes the network of stereospecific PTMs that affect the aggregation of many pathogenic agents, including Aβ. Extensive recent experimental results describe the truncated, isomerized and racemized forms of Aβ and the interplay between enzymatic and PTMs ^Sp. Currently, accumulated data suggest that non-enzymatic PTMs ^Sp occur in parallel to an existing metabolic network of enzymatic pathways, meaning that the presence and activity of enzymes does not prevent non-enzymatic reactions from occurring. PTMs ^Sp impact the functions of many proteins and peptides, including Aβ. This is in logical agreement with the silently accepted racemization hypothesis of protein aggregation (RHPA). Therefore, the ACH of AD should be complemented by the concept of PTMs ^Sp and RHPA.

Background: Some of the biochemical abnormalities underlying schizophrenia, studied in both brain and peripheral blood, involve differences in methylation and methylating enzymes, as well as other genes uncovered by chip seq or RNA seq analysis. of enzymes and some of the target genes. However, few of these studies have examined the effect of sex and drug treatment on the differences between chronic schizophrenics and controls. We present results of a larger study measuring differences in mRNA in lymphocytes of chronic schizophrenics (CSZ) and non-psychotic controls (NPC) emphasizing the differential effect os sex and antipsychotic drug treatment on the different biochemical findings.
Method(s): We studied mRNA in lymphocytes of 61 CSZ and 49 NPC subjects using qPCR assays with TaqMan probes for multiple genes to assess mRNA levels for DNMT, TET, GABAergic genes, Glucocorticoid receptor, BDNF, and several genes with high hits from RNA sequence analysis. Statistical analysis tested the effects of diagnostic status (CSZ vs. NPC) on these mRNA levels and investigated the effects of sex differences and differences in antipsychotic drug treatment on the mRNA levels in CSZ and NPC.
Result(s): DNMT1 and DNMT3A mRNAs were significantly (P < 0.01) higher only in male CSZ subjects, with the small sample of females showing no statistical difference between CSZ and NPC, but a trend in the opposite direct from the male CSZ vs NPC comparison. Several other mRNA's differences between CSZ and NPC showed a trend for a greater diagnostic difference in males than in females. The GAD1, glucocorticoid receptor (NR3C1) and CNTNPA2 mRNAs were significantly (P < 0.01) higher in CSZ than NPC. The FPRF3 mRNA was significantly (P < 0.03) lower in CSZ vs NPC, and the GAD67 mRNA showed a trend in the same direction for males (P < 0.10). In CSZ currently treated with clozapine, GABAergic mRNAs (GAD1, GAD67, GAD25) mRNA were significantly higher than in patients not treated with clozapine. CSZ treated with clozapine had significantly lower TET1 mRNA. There was a trend (P < 0.10) for NR3C1-B mRNA to be higher in clozapine patients. When we did analysis of differences between CSZ and NPC subjects, incorporating the clozapine treatment variable, it was clear that for the GABAergic mRNAs (GAD1, GAD67, GAD25), and for NR3C1-B, CNTNPA2, and IMPA2 mRNAs, that clozapine treated CSZ were primarily responsible for the difference from NPC, whereas non-clozapine treated CSZ had mRNA values more similar to NPC controls.
Conclusion(s): It is important to consider sex and antipsychotic drug treatment in comparing mRNA levels in in schizophrenic patients to controls, since some of the differences are only present in male subjects and other may be explained by clozapine treatment rather than primarily diagnosis. Many previous studies of similar differences in post-mortem brain samples, and some studies of methylation differences using peripheral blood cells, have not examined sex and drug treatment effects, and these could be possible confounds in interpreting diagnostic differences in these biochemical effects