Study on the correlation between dietary structure and sleep in patients with insomnia disorder.

OBJECTIVE: Insomnia disorder (ID) is a persistent difficulty sleeping, often accompanied by anxiety and depression, which seriously reduces a person's quality of life. Dietary changes in insomnia patients have been a concern. To explore the rationality of diet in patients with ID and its correlation with insomnia in ID patients. PATIENTS AND METHODS: This study included 216 patients diagnosed with ID and 197 individuals as the healthy control (HC) group who attended the neurology outpatient clinic or sleep clinic at Henan Provincial People's Hospital between September 2018 and November 2019. Through the Pittsburgh Sleep Quality Index (PSQI), Insomnia Severity Index (ISI), Hamilton Anxiety Scale (HAMA), and Hamilton Depression Scale (HAMD), sleep and mental conditions were assessed in the ID and HC groups. The dietary intake structure of both groups was observed using the food frequency table. Meanwhile, the relationship between dietary intake and sleep quality was analyzed based on the logistics regression. RESULTS: Individuals in the ID group had significantly higher age, weight, and body mass index compared to the HC group (p<0.01). Individuals within the ID category demonstrated a heightened daily consumption of carbohydrates, grains, tubers, and legumes relative to the healthy control group. In contrast, the intake levels of vegetables, fruits, and nuts were diminished compared to the HC group, with this difference being statistically significant (p<0.01). A positive correlation was observed between the daily consumption of grains, tubers, and legumes and PSQI scores. Conversely, a negative association was found between daily consumption of vegetables and fruits. CONCLUSIONS: ID patients exhibit an elevated intake of carbohydrates, whereas the consumption of vegetables, fruits, and nuts is deficient in comparison to the healthy cohort, implying that a distorted dietary structure might be a contributing factor to ID onset. Sensible and scientific dietary guidance is of considerable significance in preventing the onset of ID and facilitating its management. However, the derived conclusions warrant further extensive research.

[Influences of taurine and micronutrients on nitric oxide synthase expression and cGMP content in rat retina].

AIM: To investigate the influence of taurine and micronutrients on visual signal transmission. METHODS: Wistar rats were divided into three groups, that is control group, experiment group 1 and experiment group 2, and fed for 3 weeks with normal diet, 5 times and 10 times doses of requirements of taurine, vitamin A, vitamin B, zinc and selenium, then each treatment group were divided into light group and dark adaptation group. After feeding another 3 days in different environments with normal diet, all animals were killed and cGMP level and NOS expression were analysed in retina and retinogeniculate. RESULTS: The NOS expression and cGMP contents of photoreceptor cells, visual cortex and retinogeniculate were increased in dark adaptation group compared with light group. Nutritional intervention could enhance the NOS staining in dark environment, increased the cGMP contents whether light or dark condition. CONCLUSION: The distribution, expression and content of NO and cGMP are quite different in various light adaptation status. Taurine and micronutrient intervention may modurate the visual signal transmission or vision function mediated by the changes of NO or cGMP.

Myeloid cell-lineage and premylocytic-stage-specific- expression of themouse myeloperoxidase gene is controlled at initiation as well as elongation levels of transcription.

The myeloperoxidase (MPO) is an important microbicidal protein present at high concentration in the primary granule of mature granulocyte and its expression is regulated in both myeloidcell-lineage and premyelocytic-stage-specific manners. A better understanding of the underlying control mechanisms should provide insights into the temporal and co-ordinate regulation of the gene expression during granulopoiesis. We have identified its promoter by mapping the start(s) of transcription using various molecular approaches together with demonstrating the promoter function of the relevant DNA segment in a transient transfection reporter assay. Besides the major start of transcription mapped at G residue, 11 nucleotide upstream of the 3' end of exon 0, the usage of that is specific to the MPO expressing cell lines, we have shown that irrespective of the MPO-expression status of the hematopoietic cells, transcription occurs broadly within a two kb region upstream of the 5' proximity of the gene, and is largely terminated in intron 2. These data support a model of the premyelocytic-stage-specific MPO expression, the control of which is operated at initiation as well as elongation levels of transcription.

Proto-oncogene expression in a human chondrosarcoma cell line: HCS-2/8.

HCS-2/8 is a stable human chondrosarcoma cell line with many chondrocytic characteristics and has the capacity to form chondrosarcomas in nude mice. The cells display both biochemically and morphologically definable changes in sparse, subconfluent, confluent and over-confluent phases of in vitro culture. Such features of HCS-2/8 cells may reflect the processes of both proliferation and differentiation of chondrocytes in vivo. We examined the correlations of these changes of HCS-2/8 cells with their transcript levels of 21 proto-oncogenes by Northern analysis. We found no detectable transcripts of 9 proto-oncogenes (c-sis, c-met, c-src, c-lyn, c-fgr, c-ros, c-pim, Blym and N-myc), but detected transcripts of 12 other proto-oncogenes (int-2, erbB, c-abl, c-raf-1, c-fyn, K-ras, H-ras, c-mos, c-myc, c-myb, c-fos, and c-jun). In the over-confluent phase, the levels of c-fos and c-raf-1 were increased several dozen times and about 5 times, respectively, while the level of c-abl was about 1/5th of that in the sparse, subconfluent and confluent phases of culture. The level of int-2 increased about 10-fold in the confluent and over-confluent phases of in vitro culture. The transcript levels of c-mos and K-ras were high in the sparse phase, low in the subconfluent and confluent phases and high in the over-confluent phase. The levels of the other 6 proto-oncogenes in HCS-2/8 cells were constant in all phases of in vitro culture.

The myeloid-lineage specific enhancer of the mouse myeloperoxidase gene consists of three cis-elements defined as in vitro DNase I footprints.

The myeloid-lineage specific enhancer at 3.4-3.1 kb upstream of the mouse myeloperoxidase gene [1] has been further characterised. In vitro DNase I footprinting experiments revealed three protected sequences (FT-I, -II and -III) in the enhancer, associated with the proteins that are enriched in WEHI 3BD+ cells, at which the MPO gene is highly expressed; but not in two non-MPO expressing lymphocytic cell lines. Site-specific mutations at each element severely reduced the level of the reporter gene activity in a non-additive manner. This is parallel with either abolishment or alteration of the corresponding wild-type protein-DNA interaction in vitro. Consideration of the sequence motifs present in the enhancer suggests that the cis-elements defined as the in vitro DNase I footprints are likely to be novel.

[Preliminary study on RFLPs for dystrophin gene in Chinese].

We have studied the RFLPs distribution and frequency of dystrophin gene in Chinese by using 14 subclones of complete 14 kb cDNA for the dystrophin gene as hybridization probes. Allelic fragments are detected in hybridization patterns of Pvu II/1a, Taq I/2b-3, Taq I/5b-7, Xba I/10. Among them, the allelic fragments (26 kb and 3.8 kb) in Pvu II/2b-3 patterns and the allelic fragments (10 kb and 8.4 kb) in Taq I/5b-7 patterns are the new RFLPs which have never been reported. From the comparison of our data with those of Caucasian and Japanese respectively and their statistical analysis, we can obtain the results as follows: There is remarkable difference (p less than 0.01) of the allelic fragment frequency in Taq I/2 b-3 (A1 = 3.4 kb, fre. 0.04; A2 = 3.2 kb, fre. 0.96) and Xba I/10 (A1 = 7.4 kb, fre. 0.41; A2 = 6.7 kb, fre. 0.59) between Chinese and Caucasian. The frequency of the allelic fragments A2 in Taq I/8 (A1 = 6.5 kb, A2 = 5.6 kb) and EcoR V/9 (A1 = 11.8 kb, A2 = 10.7 kb) are high in Caucasian, but have not been detected in Chinese. These differences are also highly significant. But the B1B2 allelic frequencies in Taq I/5 b-7 (B1 = 3.2 kb, B2 = 1.6 kb) are the same. There is no significant difference in the frequency of the allelic fragments A1A2 and B1B2 in Pvu II/1 a between Chinese and Japanese. Preliminary results suggest that there probably are high frequencies for spontaneous neutral mutations in the evolution process of the huge dystrophin gene (about 2,300 kb). In the meantime, the neutral mutation frequencies of various sectional sequences have remarkable differences, and that of some sectional sequences of the gene between Chinese and Caucasian may also have remarkable differences.

Parallel stranded DNA under the scanning tunnelling microscope.

Using scanning tunnelling microscopy, we have directly observed parallel stranded DNA helixes of 43 nucleotides in length. The double helix is right-handed and has an average spacing, 17.43 A (+/- 1 S.D.: 2.30 A), and an average apparent depth, 4.79 A (+/- 1 S.D.: 1.04 A) for each groove. The average pitch of the helical turn is 34 A (+/- 1 S.D.: 3.35 A) and consists of no more than ten base pairs. The diameter of the helix is approx. 17-20 A. Our results provide direct evidence for the existence of a parallel structure of DNA in vitro and some details of its fine structure.

The DNA intercalator, ethidium bromide, alters the pattern of DNAse I hypersensitive sites of the beta A-globin gene in chicken erythrocytes.

We have analysed the effects of a DNA intercalator, ethidium bromide (EB), on chromatin structure in nuclei from both chicken mature erythrocytes (RBC) and reticulocytes (Ret). A differential release of nuclear proteins was obtained from both types of nuclei exposed to EB. Among these proteins, a species of 45 kDa is the major component. Furthermore, in the 10 mM EB-treated nuclei, the pattern of DNAse I hypersensitive sites (DHS) around the chicken beta A-globin gene were significantly altered, i.e., the original set was replaced by a new set of DHS. We have discussed the implications of our observations, in the light of current concepts of functional aspects of the conformational heterogeneity of DNA in both protein-DNA interactions and chromatin structure, as well as the effects of DNA intercalators on DNA conformation.

Plasmodium berghei: quantitation of in vitro effects of antimalarial drugs on exoerythrocytic development by a ribosomal RNA probe.

A stage-specific ribosomal RNA probe has been used to quantitate exoerythrocytic development of Plasmodium berghei in primary cultures of mouse hepatocytes. Parasite rRNA could be detected as soon as 6 hr after sporozoite invasion and was increased during schizogony to a maximum at 48 hr, when mature schizonts were identified by microscopy. As few as 10 exoerythrocytic schizonts could be detected by filter blot hybridization, followed by autoradiography and liquid scintillation counting. By hybridizing the culture rRNA samples with either parasite-specific or universal rRNA probes, the in vitro tissue schizonticidal activity and hepatotoxicity of primaquine, two of its analogues, and pyrimethamine, could be assessed. After a 48-hr exposure of the culture to serial dilutions of each drug, a quantitative relationship was demonstrated between the decrease of the parasite rRNA and the increase of the drug concentrations. No significant parasite-specific rRNA could be detected at the concentration achieving complete inhibition of schizont formation but causing no cytotoxic effects on host hepatocytes. In contrast to microscopic-based assays, this molecular approach provides an objective and quantitative in vitro method for rapid screening and evaluation of tissue schizonticidal antimalarials.

A Plasmodium falciparum-specific reverse target capture assay.

Plasmodium falciparum DNA is detected with an assay modeled according to the reverse target capture assay described by Morrissey et al. [19] for the detection of Listeria cells. A poly(A)-tailed oligonucleotide (pWZ34), derived from the partial sequence of a 4-kb repetitive unit of P. falciparum, functions as a capture probe and the labelled 21-bp repetitive units specific for P. falciparum serve as a reporter probe. Both probes are complementary to non-overlapping regions of the target DNA and in the presence of high concentration of chaotropic salts, hybridization efficiently takes place at relatively low temperatures (15 min. 37 degrees C). The addition of poly(dT)-derivatized ferromagnetic beads allows the formation of A:T base pairing between the tailed beads and the tailed capture probe. Upon applying magnetic force, the target-capture-reporter-probe complex attached to the beads is removed from the reaction mixture, leaving the bulk of unreacted reporter molecules behind. Subsequent washings of the immobilized complex reduces the amount of non-specifically bound reporter probe. After elution of the complex from the beads a new cycle of capture, washing and release of the target-capture-reporter-probe complex is initiated by the additions of unused (dT)-tailed beads. After 3 cycles, the signal-to-noise ratio with 0.1 pg of P. falciparum DNA as a target was as high as 21-27, with a background of 8-10 cpm. The assay is unique in its speed, well suited for large sample numbers, and allows the manipulation of the background at will by simply increasing the number of capture rounds.

Parallel stranded DNA under scanning tunnelling microscope: the main characteristics of the double helix.

Using the scanning tunnelling microscopy we have directly observed the parallel stranded DNA of 43 bp in length, containing alternating AT-stretches. The double helix is right-handed and has the same width of each grooves equal to 17.4 A. The average pitch of the helical turn is about 34 A. The parallel double helix possesses no more than 8.6 bases per one turn. The diameter of the parallel stranded DNA molecule is 17-18 A. We conclude that in parallel DNA double helix the angle between N-glycoside bounds in trans-Crick-Watson base pairs is close to 180 degrees.

Non-CS pre-erythrocytic protective antigens.

Three novel non-CS antigens have been identified on P. falciparum and P. berghei sporozoites and exoerythrocytic parasites. CSP-2 is a sporozoite surface protein common to P. falciparum and P. berghei that elicits antibody-mediated protection, and is also found within P. berghei EE parasites. LSA is a P. falciparum EE-specific antigen localized within the parasitophorous vacuole. LSA-2 is a P. berghei EE-specific antigen, localized on the parasitophorous vacuole membrane, that protected mice to P. berghei sporozoite challenge, and elicited cytotoxic T cells that killed P. berghei EE parasites in vitro.

Stage-specific ribosomal RNA expression switches during sporozoite invasion of hepatocytes.

Two structurally distinct ribosomal RNAs (rRNAs) occur in different developmental stages of malaria parasites. One point at which the transition from one type to the other is found shortly after sporozoites invade hepatocytes, the first stage of parasite development in the mammalian host. The invasion in itself appears necessary but insufficient to trigger the rRNA transition. The progression of events involved in the synthesis of a new type ribosome is tied to the fate of the invading parasite. Interestingly, the switch also occurs in irradiated sporozoites. The new rRNAs produced are processed to the mature size, indicating that rRNA transcription and processing remain normal in the attenuated parasites. These results have implications for monitoring antimalaria vaccine candidates and drug efficacy.

Pre-erythrocytic stage malaria parasites: non-circumsporozoite protein antigens.

A series of non-circumsporozoite proteins found in pre-erythrocytic parasites are being developed as putative vaccine candidates. It is anticipated that these will be useful in addition to, rather than instead of, the CS (circumsporozoite) vaccines. It is likely that a greater understanding of the basic biology of malaria parasite-host relationships will lead to development of improved malarial vaccines.

A study of the chromatin structure of human beta-like hemoglobin genes in K562 cell line with a modified assay of nick-translation of nuclei.

A modified assay of nick-translation of nuclei has been developed to study the chromatin structure of human beta-like globin genes in nuclei of K 562 cell line. Nuclei were gently digested with DNase I and nick-translated with E. coli DNA polymerase I in the presence of 32P-triphosphate nucleotides. The total DNA from the labelled nuclei was used as probes to hybridize restricted fragments of beta-like globin genes which have been immobilized on Diazobenzyloxymethyl (DBM) paper. Using this approach we have observed that in K 562 nuclei all beta-like globin genes, including epsilon, gamma, delta, and beta-globin genes and human 18 S ribosomal genes are preferentially labelled in comparison to alpha-lactalbumin and c-sis genes which do not express in K 562 cells, but the total DNA from nick-translated nuclei of a nonerythroid cell line hybridized none of those genes except for 18 S ribosomal gene. This assay is a simple and fast method for surveying chromatin structure of any individual DNA sequence in nuclei once the corresponding clone is available.

Antisense RNA: effect of ribosome binding sites, target location, size, and concentration on the translation of specific mRNA molecules.

A series of plasmids were constructed to generate RNA complementary to the beta-galactosidase messenger RNA under control of the phage lambda PL promoter. These plasmids generate anti-lacZ mRNA bearing or lacking a synthetic ribosome binding site adjacent to the lambda PL promoter and/or the lacZ ribosome binding site in reverse orientation. Fragments of lacZ DNA from the 5' and/or the 3' region were used in these constructions. When these anti-mRNA molecules were produced in Escherichia coli 294, maximal inhibition of beta-galactosidase synthesis occurred when a functional ribosome binding site was present near the 5' end of the anti-mRNA and the anti-mRNA synthesized was complementary to the 5' region of the mRNA corresponding to the lacZ ribosome binding site and/or the 5'-coding sequence. Anti-mRNAs producing maximal inhibition of beta-galactosidase synthesis exhibited an anti-lacZ mRNA:normal lacZ mRNA ratio of 100:1 or higher. Those showing lower levels of inhibition exhibited much lower anti-lacZ mRNA:normal lacZ mRNA ratios. A functional ribosome binding site at the 5'-end was found to decrease the decay rate of the anti-lacZ mRNAs. In addition, the incorporation of a transcription terminator just downstream of the antisense segment provided for more efficient inhibition of lacZ mRNA translation due to synthesis of smaller and more abundant anti-lacZ mRNAs. The optimal constructions produced undetectable levels of beta-galactosidase synthesis.