[A multicenter study on respiratory pathogen detection with Mycoplasma pneumoniae pneumonia in children].

Objective: To analyze the status of respiratory pathogen detection and the clinical features in children with Mycoplasma pneumoniae pneumonia (MPP). Methods: A prospective, multicenter study was conducted to collect clinical data, including medical history, laboratory examinations and multiplex PCR tests of children diagnosed with MPP from 4 hospitals in China between November 15th and December 20th, 2023. The multiplex PCR results and clinical characteristics of MPP children in different regions were analyzed. The children were divided into severe and mild groups according to the severity of the disease. Patients in the severe group were further divided into Mycoplasma pneumoniae (MP) alone and Multi-pathogen co-detection groups based on whether other pathogens were detected besides MP, to analyze the influence of respiratory pathogen co-detection rate on the severity of the disease. Mann-Whitney rank sum test and Chi-square test were used to compare data between independent groups. Results: A total of 298 children, 136 males and 162 females, were enrolled in this study, including 204 children in the severe group with an onset age of 7.0 (6.0, 8.0) years, and 94 children in the mild group with an onset age of 6.5 (4.0, 7.8) years. The level of C-reactive protein, D-dimer, lactic dehydrogenase (LDH) were significantly higher (10.0 (5.0, 18.0) vs. 5.0 (5.0, 7.5) mg/L, 0.6 (0.4, 1.1) vs. 0.5 (0.3, 0.6) mg/L, 337 (286, 431) vs. 314 (271, 393) U/L, Z=2.02, 2.50, 3.05, all P<0.05), and the length of hospitalization was significantly longer in the severe group compared with those in mild group (6.0 (6.0, 7.0) vs. 5.0 (4.0, 6.0) d, Z=4.37, P<0.05). The time from onset to admission in severe MPP children was significantly shorter than that in mild MPP children (6.0 (5.0, 9.5) vs. 9.0 (7.0, 13.0) d, Z=2.23, P=0.026). All patients completed the multiplex PCR test, with 142 cases (47.7%) MPP children detected with 21 pathogens including adenovirus 25 cases (8.4%), human coronavirus 23 cases (7.7%), rhinovirus 21 cases (7.0%), Streptococcus pneumoniae 21 cases (7.0%), influenza A virus 18 cases (6.0%). The pathogens with the highest detection rates in Tianjin, Shanghai, Wenzhou and Chengdu were Staphylococcus aureus at 10.7% (8/75), adenovirus at 13.0% (10/77), adenovirus at 15.3% (9/59), and both rhinovirus and Haemophilus influenzae at 11.5% (10/87) each. The multi-pathogen co-detection rate in severe MPP children was significantly higher than that in mild MPP group (52.9% (108/204) vs. 36.2% (34/94), χ²=10.62,P=0.005). Among severe MPP children, there are 89 cases in the multi-pathogen co-detection group and 73 cases in the simple MPP group. The levels of LDH, D-dimer and neutrophil counts in the multi-pathogen co-detection group were significantly higher than those in the simple MPP group (348 (284, 422) vs. 307 (270, 358) U/L, 0.8 (0.5, 1.5) vs. 0.6 (0.4, 1.0) mg/L, 4.99 (3.66, 6.89)×109 vs. 4.06 (2.91, 5.65)×109/L, Z=5.17, 4.99, 6.11, all P<0.05). Conclusions: The co-detection rate of respiratory pathogens, LDH and D-dimer in children with severe MPP were higher than those with mild MPP. Among severe MPP children the stress response of children in co-detection group was more serious than that of children with simple MPP.

[Epidemiological investigation on a visceral leishmaniasis case in Zhengzhou City of Henan Province].

OBJECTIVE: To perform an epidemiological investigation on a case with visceral leishmaniasis in Zhengzhou City, Henan Province, and to identify the source of infection, so as to illustrate the transmission chain and assess the risk of local leishmaniasis transmission. METHODS: The medical data were collected from a case with visceral leishmaniasis in Zhengzhou City, and the patient's bone marrow smears were detected by microscopy. Serum anti-Leishmania antibody test and PCR assay were performed among high-risk residents and all dogs in the village where the patient lived. Sandflies were captured using light traps and artificial traps, and the captured female Phlebotomus chinensis was subjected to PCR assay. The internal transcribed spacer 1 (ITS1) gene was amplified with a nested PCR assay using the genomic DNA extracted from visceral leishmaniasis patients, positive dogs and sandflies, and the sequences were aligned with those download from NCBI. In addition, a phylogenetic tree was created based on the ITS1 gene. RESULTS: The visceral leishmaniasis patient had recurrent irregular fever, reduced complete blood counts, low hemoglobin, and a large number of Leishmania amastigotes in bone marrow smears, and was therefore diagnosed as visceral leishmaniasis. Both rk39 rapid diagnostic test and PCR assay tested negative among 324 residents living neighboring the patient's residence, while 21.39% (43/201) dogs were positive for rk39 rapid diagnostic test and 13.93% (28/201) positive for PCR assay. There were 17 female Ph. chinensis tested positive for Leishmania (0.82%) by PCR assay, and the ITS gene sequences from visceral leishmaniasis patients, positive dogs and sandflies shared a 100% homology with L. infantum. The Leishmania species was therefore characterized as L. infantum. CONCLUSIONS: L. infantum infection occurs in visceral leishmaniasis patients, dogs and sandflies in Zhengzhou City, indicating a complete transmission chain and a high transmission risk of visceral leishmaniasis by L. infantum. Intensified control measures are required to prevent local transmission of leishmaniasis in Zhengzhou City.

[Susceptibility of Anopheles sinensis to insecticides in Puyang City, Henan Province].

OBJECTIVE: To investigate the susceptibility of Anopheles sinensis to malathion, deltamethrin and lambda-cyhalothrin in Puyang City, Henan Province, so as to provide the scientific basis for local malaria vector control. METHODS: An. sinensis was captured from Puyang County, Puyang City of Henan Province in September 2018 and July 2020, and the susceptibility of field captured An. sinensis to malathion, deltamethrin and lambda-cyhalothrin was tested using the filter-paper bioassay recommended by WHO. The insecticide resistance level was assessed based on the WHO criteria. RESULTS: In 2018 and 2010, the half knock-down times (KT50) of malathion were 91.08 min and 40.95 min for An. sinensis, with knock-down rates of 37.50% and 60.87% 60 min post-exposure to malathion and 24-hour mortality rates of 90.91% and 100%, respectively, and the insecticide resistance levels were moderately resistant (M) and susceptible (S). The KT50 of deltamethrin were 415.56 min and 341.19 min for An. sinensis in 2018 and 2020, with knock-down rates of 22.92% and 16.98% 60 min post-exposure to malathion and 24-hour mortality rates of 22.92% and 16.98%, and the insecticide resistance levels were all resistant (R). The KT50 of lambda-cyhalothrin were 164.22 min and 236.22 min for An. sinensis in 2018 and 2020, with knock-down rates of 30.39% and 38.30% 60 min postexposure to malathion and 24 h mortality rates of 19.60% and 21.28%, respectively, and the insecticide resistance levels were all R. CONCLUSIONS: An. sinensis is relatively susceptible to malathion but has developed high-level resistance to deltamethrin and lambda-cyhalothrin in Puyang City, Henan Province..

Study on the effects of quercetin on brain cell apoptosis and HMGB-1 and TLR-4 level in rats with encephalitis.

OBJECTIVE: To explore the therapeutic effects of quercetin on rats with encephalitis, especially on cell apoptosis, and the levels of HMGB-1 and TLR-4. MATERIALS AND METHODS: 32 healthy rats were equally assigned into ZC group (healthy group), NY group (encephalitis group), DJ group (60 ml quercetin group), and GJ group (240 ml quercetin group) followed by analysis of cell apoptosis, brain tissue water content, neurons, HMGB-1, TLR-4 and other inflammatory factors. RESULTS: The ZC group showed normal neuron volume and equitable staining; compared with ZC group, NY group showed neuron volume shrinkage and chromatin condensation; the neuron and color in DJ group were slightly better than NY group; the neuron volume in GJ group increased significantly and chromatin is distributed evenly. TLR-4, IL-4, IL-6, HMGB-1 in ZC and DJ group were significantly lower than those in NY group (p<0.05); IL-4, IL-6, HMGB-1 in GJ group significantly decreased compared with DJ group (p<0.05); MMP-9 enzyme activity in ZC and DJ group was significantly lower than NY group (p<0.05) with lower level in GJ group than DJ group (p<0.05). The water content was higher in brain tissue of NY group than ZC group (p<0.05) and lower in DJ group than NY group (p<0.05) with lower level in GJ group than DJ and NY group (all p<0.05). The hippocampal neurons and cortical neurons in ZC and DJ group were higher than those in NY group (p<0.05) and elevated in GJ group compared with DJ group and NY group (p<0.05). CONCLUSIONS: Quercetin is effective in treating encephalitis rats possibly through inhibition of neuronal cell apoptosis and level of HMGB-1 and TLR-4.

[Expression and clinical significance of CXC chemokine ligand 10 in glioma].

Objective: To analyze the expression of CXC chemokine ligand 10 (CXCL10) in glioma and its clinical significance through bioinformatics. Methods: The expression level of CXCL10 in glioma, and its prognostic significance, gene ontology (GO) function annotation, Kyoto Encyclopedia of Genes and Genome (KEGG) pathway enrichment and the correlation of tumor cell purity were analyzed in TCGA, CGGA, MetaScape, TIMER databases. In addition, 34 clinical glioma tissues were collected for Western Blot and immunohistochemistry to further verify the correlation between CXCL10 and glioma. Results: CGGA and TCGA database analysis showed that with the increase of WHO grade, the expression of CXCL10 in gliomas increased (P<0.01). The overall survival rate of patients with high CXCL10 expression was significantly lower than that of patients with low expression (χ2 =148.1,P<0.05). Among patients with grade Ⅳ glioblastoma who received radiotherapy or chemotherapy, the patients with low CXCL10 expression were associated with good survival (χ2 =6.714,P<0.05;χ2 =5.618,P<0.05). Moreover, GO and KEGG analysis showed that genes co-expressed with CXCL10 were mainly enriched in the biological processes such as cytokine-mediated signaling pathways, regulating adaptive immune responses and inflammatory responses. Furthermore, TIMER database analysis showed that CXCL10 was negatively correlated with the purity of glioma cells (LGG: r=-0.129;GBM: r=-0.165;P<0.05). Similarly, clinical sample analysis also showed that the expression level of CXCL10 increased in glioma, and it increased with the grade of glioma (all P<0.05). Conclusion: The expression of CXCL10 is up-regulated in glioma as well as it increased with the malignant degree of glioma. At the same time, the high expression of CXCL10 in glioma is closely related to the poor prognosis of patients.

[Epidemic status of malaria and progress of malaria elimination in Henan Province, 2018].

OBJECTIVE: To understand the epidemic status of malaria and progress of malaria elimination in Henan Province in 2018, so as to provide the scientific evidence for formulating the control strategy of malaria. METHODS: All data pertaining to the epidemic status of malaria and malaria case were captured from Henan Province in 2018, and the epidemic status of malaria and the diagnosis and treatment of malaria cases were analyzed using a descriptive epidemiological method. RESULTS: A total of 174 malaria cases were reported in Henan Province in 2018, with a male-to-female ratio of 33.8∶1. The cases were predominantly found at 30- and 40- years, and farmer was the highest-risk population. All cases were imported for overseas countries and 96.55% (168/174) were from Africa. The cases were reported across 17 cities of the province, and 63.22% (110 cases) were detected in 4 cities of Zhengzhou, Luoyang, Xuchang and Puyang. Both the median duration from onset to initial admission and the median time from the initial admission to definitive diagnosis were 1 d. There were 6 cases (3.45%) with more than 7 days to visit a doctor, and there were 13 cases (7.47%) with definitive diagnosis of over 7 days (delay in definitive diagnosis). All malaria cases were reported within 24 h, and 98.28% (171/174) completed case investigations within 3 d. All 18 cities had achieved malaria elimination by December 2018. CONCLUSIONS: All malaria cases reported in Henan Province in 2018 are imported from overseas countries. Intensifying malaria surveillance among returnees from Africa and Southeast Asia and timely identification and treatment of imported malaria cases are required to prevent the development of secondary malaria cases, so as to ensure the achievement of malaria elimination in Henan Province on schedule.

[Epidemiological characteristics of malaria in Henan Province from 1950 to 2019].

OBJECTIVE: To analyze the epidemiological characteristics of malaria in Henan Province from 1950 to 2019, so as to provide the scientific evidence for consolidating malaria elimination achievements in the province. METHODS: The epidemiological situation of malaria and demographic data in Henan Province from 1950 to 2019 were collected, and the statistical analyses were performed using a descriptive epidemiological method. The spatial temporal distribution of malaria cases was identified using the software ArcGIS version 10.3. RESULTS: During the period from 1950 through 2019, the progress of malaria elimination was divided into 4 stages in Henan Province, including the baseline-survey and key-control stage, morbidity-control and incidence-reduction stage, basic-eradication and achievement-consolidation stage and elimination stage. The spatial distribution of malaria cases shifted from south of the Huai River and the plain regions between the Yellow River and Taihang Mountain to the Huang-Huai-Hai Plain and Nanyang Basin, then was concentrated in eastern part of southern Huai River where Anopheles anthropophagus was distributed, and finally was gradually under control following malaria outbreak in Eastern Henan Plain. In addition, the species of Plasmodium changed from P. vivax, P. falciparum and P. malariae co-endemics to a single P. vivax infection, and the current co-endemics of 5 invasive malaria parasites, and the malaria vectors shifted from co-existence of Anopheles sinensis and An. anthropophagus to An. sinensis alone. CONCLUSIONS: There has been a large change in the epidemiological characteristics of malaria in Henan Province from 1950 to 2019. Although malaria has been eliminated in Henan Province, the consolidation of the malaria elimination achievements remain a great challenge due to overseas imported malaria.

[The first identification of epidemic clone of enterotoxic Escherichia coli O∶6 serogroup highly associated with azithromycin resistance in Shanghai].

Objective: To investigate the molecular characterization of adult diarrhea cases caused by enterotoxic Escherichia coli (ETEC) and explore the practical model of epidemiology for laboratory technique and data needs based on the surveillance network. Methods: Epidemiological design and sampling targeted adult cases ETEC caused diarrhea in epidemic season. The enterotoxin type, serogroup, resistance, colonization factor and molecular type of ETEC were identified. Multiple dynamic phenotypic characteristics of ETEC were indicated by multidimensional and multivariable data. Results: From 2016 to 2018, 84 eligible ETEC strains were detected. The dominant serums/toxins were O∶6 (STh), O∶25 (LT), O∶159 (STh), O∶153 (STh). O∶6 (STh+CS21), which replaced O∶25 and O∶159 as the popular clones in 2018. Six cases of O∶153 (STh+CFA/I+CS8+PT34) in outbreak in 2017 were imported ones. The resistance rates of ETEC strains detected in adults to sulfasoxazole, naproxinic acid, ampicillin and azithromycin were more than 30%, multidrug resistance (MDR) reached 58.3%. Serum/toxin types suggested that attenuated strains were more likely to become MDR. Molecular typing confirmed that the genetic similarity of the dominant clone of O∶6 serogroup (PT20-24) was higher than O∶25 and O∶159. There was a high correlation between the minimal inhibitory concentration (MIC) of azithromycin and the resistant gene mphA (87.5%, 28/32). O∶6 (STh+CS21+mphA) resistant clone was first detected in 2016. Conclusion: A new epidemic clone in adult ETEC diarrhea cases in Shanghai was O∶6 (STh+CS21+mphA). For the first time the association between azithromycin resistance gene mphA and a serum group of ETEC was observed. Multidimensional and multivariate analysis techniques based on epidemiology can help reveal the potential transmission pattern of ETEC for the accurate surveillance and early warning of outbreaks.

[Stratified sampling survey of major human parasitic diseases in Henan province].

Objective: To understand the prevalence of major human parasitic diseases and related factors in Henan province. Methods: This stratified sampling survey was carried out according to the requirement of national survey protocol of major human parasitic diseases, 2014-2015. The prevalence of soil-transmitted helminths infection, taeniasis and intestinal protozoiasis were surveyed in 104 sites selected from 35 counties (districts) and the prevalence of clonorchiasis was surveyed in 62 sites selected from 37 townships. In each survey spot, 250 persons were surveyed. A total of 26 866 persons and 15 893 persons were surveyed. Modified Kato-Katz thick smear was used to detect the eggs of intestinal helminthes. Tube fecal culture was used to identify the species of hookworm. The Enterobius eggs were detected in children aged 3 to 6 years by using adhesive tape. The cyst and trophozoite of intestinal protozoa were examined with physiological saline direct smear method and iodine stain method. Results: The overall infestation rate of intestinal parasites was2.02% in Henan, and the worm infection rate was higher than protozoa infection rate. Fourteen kinds of intestinal parasites were found, including nematode (5 species), trematode (2 species), and protozoan (7 species). The infection rate of Enterobius vermicularis was highest, and Qinba Mountain ecological area had the highest infestation rate of intestinal parasites in 4 ecological areas of Henan. There was no significant difference in intestinal parasite infection rate between males and females (χ(2)=3.630, P=0.057), and the differences in intestinal parasite infection rate among different age groups had significance (χ(2)=124.783, P=0.000 1). The infection rate reached the peak in age group ≤9 years and the major parasite was Enterobius vermicularis. Furthermore the overall human infection rate of parasite showed a downward trend with the increase of educational level of the people (χ(2)=70.969, P=0.000 1), the differences had significance (χ(2)=120.118, P=0.000 1). For different populations, the infection rate of intestinal parasites was highest among preschool children. The infection of intestinal helminth was mainly mild, only 2 severe cases were detected. The infection rate of Clonorchis sinensis in urban residents was only 0.006%. Logistic regression analysis showed that being preschool children (χ(2)=15.765, P=0.000 1) and drinking well water (χ(2)=45.589, P=0.000 1) were the risk factors for intestinal parasite infection, and annual income per capita of farmers was the protective factor against intestinal parasite infection. The infection rates of protozoa and intestinal parasites decreased sharply compared with the results of previous two surveys, and the rate of intestinal helminth infection also dropped sharply compared with the second survey. The numbers of protozoa, helminth and intestinal parasites detected in this survey were all less than the numbers found in the previous two surveys. Conclusions: Compared the results of three surveys in Henan, the infection rate of protozoa and intestinal parasites showed a downward trend. The prevention and treatment of Enterobius vermicularis infection in children should be the key point of parasitic disease control in the future.

Mean platelet volume, platelet distribution width and platelet count in erectile dysfunction: A systematic review and meta-analysis.

The aim of this study was to investigate the relationship between mean platelet volume (MPV), platelet distribution width (PDW), platelet count (PC) and erectile dysfunction (ED). We searched for observational studies from PubMed, EMBASE, Web of Science and CNKI up to 31 March 2016. Two reviewers independently selected the studies and extracted the data. MPV, PDW, and PC and mean differences in these platelet indices between healthy subjects and ED patients were explored using the Comprehensive Meta-Analysis software package. Seven studies including 795 patients and 524 healthy subjects met the inclusion criteria. The MPV was significantly larger in patients with ED than controls with the standardised mean difference of 0.596 fL (95% CI: 0.378, 0.815, p < 0.001). In ED patients, the pooled mean difference in MPV between vasculogenic ED patients and nonvasculogenic ED patients was 0.706 fL in case-control studies (95% CI: 0.410, 1.002, p < 0.001). There was no significant difference in PDW and PC between healthy subjects and ED patients. The available data suggest that larger MPV was associated with ED. Patients with vasculogenic ED tend to have higher MPV than nonvasculogenic ED patients. Further studies are needed to assess whether increased MPV in ED patients is associated with increased cardiovascular disease.

[Clinical evaluation value of visual analogue scale combined with C-reactive protein for the prognosis of peritonsillar abscess].

Objective:The aim of this study is to investigate the clinical value of visual analogue scale combined with serum C-reactive protein in evaluating the prognosis of peritonsillar abscess. Method:Thirty-one patients be hospitalized with peritonsillar abscess were enrolled in this study, who were treated by puncture and antibiotic from January 2016 to February 2017. VAS and CRP level were detected on the 1, 3, 5 day of hospitalization for all the patients. Result:Twenty-five patients were cured after 5 days. The 1, 3, 5 day's VAS of forty-seven patients were 8.20±1.25, 4.42±1.05, 1.41±0.55, respectively, and the CRP level were (62.41±8.61), (20.46±5.32), (5.41±1.95)mg/L, respectively (P<0.05) in the three days. The 1, 3 day's VAS of six patients were 8.41±1.44, 7.37±1.15, respectively, and the CRP level were (59.85±8.35), (45.33±7.46)mg/L, with no significant difference (P>0.05) in the two days. But the fifth day' VAS was 1.55±0.65, and the CRP level was 10.24±2.57 mg/L, with statistically significant difference (P<0.05)) compared with the first day. Conclusion:Subjective evaluation used by VAS and objective evaluation used by CRP level to assess the prognosis of peritonsillar. Therefore, VAS score combined with C-reactive protein detection is more accurate for the prognosis of tonsil abscess evaluation..

The dependence of extracted current on discharge gas pressure in neutral beam ion sources on HL-2A tokamak.

The discharge gas pressure is a key factor to influence the extracted current of ion source. In this paper, the dependence of extracted current on discharge gas pressure was investigated in detail at different arc discharge currents. The discharge gas pressure with a very broad range (0.1 Pa-2.7 Pa) was scanned for the first time. It is turned out that, with the increasing of discharge gas pressure, the extracted current increases and the arc voltage decreases at different arc currents; however, when the discharge gas pressure exceeds a certain value, the extracted current decreases. For the same discharge gas pressure, the higher the arc current, the higher the arc voltage and the extracted current are. The arc efficiency was also calculated, and its dependence on gas pressure was almost the same with the dependence of extracted current on gas pressure, but at the same discharge gas pressure, the lower the arc current, the higher the arc efficiency is and the lower the extracted current is.

Salivary anticandidal activity and saliva composition in an HIV-infected cohort.

This study investigated salivary anticandidal activity and salivary composition in stimulated whole saliva of 18 advanced HIV-infected patients and compared these values to healthy controls. Stimulated whole saliva from HIV-infected patients showed decreased anticandidal activity. The flow rate was reduced by 40% as compared with controls. The saliva flow rate for HIV-infected patients who had recoverable yeast in their saliva was reduced as compared to HIV-infected patients without recoverable yeast. For HIV-infected patients, the saliva concentrations of lactoferrin, secretory IgA and Cl- were increased while the secretion rate of lysozyme, total protein and K+ were reduced. There was no difference in any parameter as a function of taking the antifungal drug fluconazole. There was no association between salivary anticandidal activity and any salivary component. This study shows reduced anticandidal activity and salivary flow rate in HIV-infected patients. These alterations may contribute to their increased incidence of oral candidal infections.

3' untranslated region-mediated regulation of luteinizing hormone/human chorionic gonadotropin receptor expression.

Multiple luteinizing hormone/human chorionic gonadotropin (LH/hCG) receptor mRNAs (6.7, 4.4, 2.6, and 1.8 kb) have been identified in the rat ovary. Our laboratory has previously cloned a 3.5 kb cDNA that corresponds to the 3' untranslated region (3' UTR) of the 6.7 kb transcript, the major LH/hCG receptor mRNA in rat ovary. In order to determine the effects of the 3' UTR on receptor expression, we have constructed cDNAs corresponding to the open reading frame of LH/hCG receptor or luciferase, plus these constructs with the addition of the 3' UTRs associated with the short (4.4 kb) and long (6.7 kb) LH/hCG receptor transcripts, and measured receptor or luciferase expression in 293 cells transformed with large T antigen (293T). Ligand binding analysis with 125I-hCG revealed that the 3' UTR inhibited receptor expression, which occurs through posttranscriptional events. First, the 3' UTRs reduced receptor mRNA half-life in actinomycin D-arrested cells, as compared to the open reading frame alone. Second, LH/hCG receptor mRNAs with the long 3' UTR associated with significantly fewer ribosomes. The effect of the LH/hCG receptor 3' UTRs on luciferase expression was also determined. The short 3' UTR increased luciferase activity, whereas the long 3' UTR decreased luciferase expression. Thus, the short 3' UTR exerts opposite effects on receptor and luciferase expression. However, sequences in the long 3' UTR are sufficient to inhibit both receptor and luciferase expression in 293T cells.

Molecular cloning of a novel luteinizing-hormone/human-chorionic-gonadotropin-receptor cDNA. Identification of a long 3' untranslated region and cDNA sequence of the major transcript in rat ovary.

The biological action of luteinizing hormone/human chorionic gonadotropin (lutropin/choriogonadotropin) in the ovary is mediated by interaction with its specific receptor. Lutropin/choriogonadotropin-receptor hnRNA is processed into multiple mRNAs. However, nucleotide sequences for many of the transcripts, including the major form (6.7 kb), have yet to be determined. In an attempt to identify a cDNA encoding the major transcript, we have isolated a 3.5-kb cDNA clone from a rat ovary cDNA library. The 3.5-kb cDNA recognized only two (6.7 kb, 4.4 kb) of the three (6.7, 4.4, 2.6 kb) ovarian lutropin/choriogonadotropin-receptor transcripts when used as a probe. The first 732 nucleotides of the newly identified 3.5-kb cDNA showed 98% identity to the 3' untranslated region (3' UTR) of the previously cloned cDNA corresponding to the 4.4-kb transcript. Southern blot analysis indicated that the 3.5-kb cDNA and the C-terminal domain of the lutropin/choriogonadotropin-receptor originate from the same gene. Oligonucleotide-directed cleavage of the 6.7-kb lutropin/choriogonadotropin-receptor mRNA by RNase H revealed that the newly identified 3.5-kb cDNA is a 3' extension of the 4.4-kb transcript. We propose that the nucleotide sequence of the 6.7-kb lutropin/choriogonadotropin-receptor transcript, the major form found in rat ovary, contains a long 3' UTR, which has not been previously identified.

Loss of lutropin/human choriogonadotropin receptor messenger ribonucleic acid during ligand-induced down-regulation occurs post transcriptionally.

Previous studies have demonstrated that ligand-induced down-regulation of the LH/hCG receptor in rat corpus luteum results in a loss of ligand-binding activity and a parallel decline in steady state levels of receptor mRNA. The purpose of the present study was to determine whether this loss of receptor mRNA during receptor down-regulation is due to inhibition of transcription or increased degradation of the receptor mRNA. To differentiate between these two possibilities, nuclear run-off assays were performed to study transcription rates of nuclei isolated from control and hCG down-regulated rat ovaries. A 750-mer LH/hCG receptor cDNA probe, spanning the carboxy-terminal region of the peptide and the 3'-untranslated region of the receptor cDNA, was constructed by polymerase chain reaction. RNA transcripts were synthesized from existing nuclear RNA from control and down-regulated nuclei and hybridized to the receptor cDNA probe immobilized on nylon membranes and subjected to autoradiography. Hybridization to actin cDNA was also run alongside as a control. The results of run-off transcription analysis indicated that during down-regulation, the transcription rates of LH/hCG receptor mRNA are not decreased compared to those in saline-treated controls. Although no decrease in the transcription rates of the receptor mRNA was seen in the down-regulated state, the steady state levels of the receptor mRNA showed a decline when assayed by either solution hybridization or Northern blot analysis. Furthermore, hCG induced down-regulation appears to increase the transcriptional activity of the nuclei. Estimates of steady state LH/hCG receptor mRNA turnover rates indicate that the half-life of the message in down-regulated ovaries is markedly reduced compared to that in the control. It is concluded that the loss of steady state LH/hCG receptor mRNA is not due to a decrease in transcription, but probably represents an increased degradation of the receptor mRNA.