[The role and mechanism of tumor metastasis-associated gene 1 in radiosensitivity of HeLa cells].

Objective: To determine the effect of tumor metastasis-associated gene 1 (MTA1) on the sensitivity of HeLa cells to radiotherapy, and to clarify its molecular mechanism. Methods: The transcriptome differences between MTA1 knocked down Hela cells and control cells were analyzed, and the differentially expressed genes (DEGs) was used to perform Gene-Set Enrichment Analysis (GSEA) and Gene Ontology (GO) cluster analysis. Flow cytometry was used to detect apoptosis in MTA1-overexpressed HeLa cells and control cells before and after 10 Gy X-ray irradiation. Cloning formation assay and real-time cellular analysis (RTCA) were used to monitor the cell proliferation before and after 2 Gy X-ray irradiation. To dissect the underlying molecular mechanisms of MTA1 affecting the sensitivity of radiotherapy, the proteins encoded by the DEGs were selected to construct a protein-protein interaction network, the expression of γ-H2AX was detected by immunofluorescence assay, and the expression levels of γ-H2AX, ß-CHK2, PARP and cleaved caspase 3 were measured by western blot. Results: By transcriptome sequencing analysis, we obtained 649 DEGs, of which 402 genes were up-regulated in MTA1 knockdown HeLa cells and 247 genes were down-regulated. GSEA results showed that DEGs associated with MTA1 were significantly enriched in cellular responses to DNA damage repair processes. The results of flow cytometry showed that the apoptosis rate of MTA1 over-expression group (15.67±0.81)% after 10 Gy X-ray irradiation was significantly lower than that of the control group [(40.27±2.73)%, P<0.001]. After 2 Gy X-ray irradiation, the proliferation capacity of HeLa cells overexpressing MTA1 was higher than that of control cells (P=0.024). The numbers of colon in MTA1 over-expression group before and after 2 Gy X-ray irradiation were (176±7) and (137±7) respectively, higher than (134±4) and (75±4) in control HeLa cells (P<0.05). The results of immunofluorescence assay showed that there was no significant expression of γ-H2AX in MTA1 overexpressed and control HeLa cells without X-ray irradiation. Western blot results showed that the expression level of ß-CHK2 in MTA1-overexpressing HeLa cells (1.04±0.06) was higher than that in control HeLa cells (0.58±0.25, P=0.036) after 10 Gy X-ray irradiation. The expression levels of γ-H2AX, PARP, and cleaved caspase 3 were 0.52±0.13, 0.52±0.22, and 0.63±0.18, respectively, in HeLa cells overexpressing MTA1, which were lower than 0.87±0.06, 0.78±0.12 and 0.90±0.12 in control cells (P>0.05). Conclusions: This study showed that MTA1 is significantly associated with radiosensitivity in cervical cancer HeLa cells. MTA1 over-expression obviously reduces the sensitivity of cervical cancer cells to X-ray irradiation. Mechanism studies initially indicate that MTA1 reduces the radiosensitivity of cervical cancer cells by inhibiting cleaved caspase 3 to suppress apoptosis and increasing ß-CHK2 to promote DNA repair.

[Impact of bicuspid aortic valve or tricuspid aortic valve on hemodynamics and left ventricular reverse remodeling in patients after transcatheter aortic valve replacement].

Objective: To compare the impact of bicuspid aortic valve (BAV) or tricuspid aortic valve (TAV) on hemodynamics and left ventricular reverse remodeling after transcatheter aortic valve replacement (TAVR). Methods: We retrospectively analyzed the clinical data of patients who underwent TAVR in our hospital from January 2019 to March 2021. Patients were divided into BAV group and TAV group according to aortic contrast-enhanced CT. Each patient was followed up by N-terminal pro B-type natriuretic peptide (NT-proBNP) and echocardiography at four time points, namely before TAVR, 24 hours, 1 month and 6 months after TAVR. Echocardiographic data, including mean pressure gradient (MPG), aortic valve area (AVA), left ventricular ejection fraction (LVEF), left ventricle mass (LVM) and LV mass index (LVMi) were evaluated. Results: A total of 41 patients were included. The age was (75.0±8.6) years, and male patients accounted for 53.7%. There were 19 BAV patients and 22 TAV patients in this cohort. All patients undergoing TAVR using a self-expandable prosthesis Venus-A valve. MPG was (54.16±21.22) mmHg(1 mmHg=0.133 kPa) before TAVR, (21.11±9.04) mmHg at 24 hours after TAVR, (18.84±7.37) mmHg at 1 month after TAVR, (17.68±6.04) mmHg at 6 months after TAVR in BAV group. LVEF was (50.42±13.30)% before TAVR, (53.84±10.59)% at 24 hours after TAVR, (55.68±8.71)% at 1 month after TAVR and (57.42±7.78)% at 6 months after TAVR in BAV group. MPG and LVEF substantially improved at each time point after operation, and the difference was statistically significant (all P<0.05) in BAV group. MPG in TAV group improved at each time point after operation, and the difference was statistically significant (all P<0.05). LVMi was (164.13±49.53), (156.37±39.11), (146.65±38.84) and (134.13±39.83) g/m2 at the 4 time points and the value was significantly reduced at 1 and 6 months post TAVR compared to preoperative level(both P<0.05). LVEF in the TAV group remained unchanged at 24 hours after operation, but it was improved at 1 month and 6 months after operation, and the difference was statistically significant (all P<0.05). LVMi in TAV group substantially improved at each time point after operation, and the difference was statistically significant (all P<0.05). NT-proBNP in both two groups improved after operation, at 1 month and 6 months after operation, and the difference was statistically significant (all P<0.05). MPG in TAV group improved better than in BAV group during the postoperative follow-up (24 hours after TAVR: (11.68±5.09) mmHg vs. (21.11±9.04) mmHg, P<0.001, 1 month after TAVR: (10.82±3.71) mmHg vs. (18.84±7.37) mmHg, P<0.001, 6 months after TAVR: (12.36±4.42) mmHg vs. (17.68±6.04) mmHg, P=0.003). There was no significant difference in NT-proBNP between BAV group and TAV group at each time point after operation (all P>0.05). There was no significant difference in paravalvular regurgitation and second prosthesis implantation between the two groups (all P>0.05). Conclusions: AS patients with BAV or TAV experience hemodynamic improvement and obvious left ventricular reverse remodeling after TAVR, and the therapeutic effects of TAVR are similar between BAV and TAV AS patients in the short-term post TAVR.

The circular RNA circPTK2 inhibits EMT in hepatocellular carcinoma by acting as a ceRNA and sponging miR-92a to upregulate E-cadherin.

OBJECTIVE: Hepatocellular carcinoma (HCC) is a common malignant tumor. Increasing evidence has demonstrated that microRNAs (miRNAs) play an important role in a wide variety of cellular processes. However, there are few reports about the role and underlying molecular mechanisms of miRNAs in HCC. PATIENTS AND METHODS: qRT-PCR and Western blots were performed to quantify the expression of miR-92a, E-cadherin, and circPTK2. Proliferation and invasion assays were performed to explore the function of miR-92a and circPTK2. A Luciferase assay was used to test the relationship between miR-92a, E-cadherin, and circPTK2. RESULTS: In this study, we found that miR-92a was upregulated in HCC tissues and HCC cell lines. Overexpression of miR-92a enhanced cell proliferation and invasion by targeting the E-cadherin 3'UTR in HCC cells. Furthermore, we found that circPTK2 inhibited EMT by inhibiting miR-92a, preventing its ability to downregulate E-cadherin in HCC cells. CONCLUSIONS: We identified a regulatory axis comprising circPTK2/miR-92a/E-cadherin in HCC cells that may serve as a valuable biomarker and therapeutic target for patients with HCC.

Changes in distribution of labile zinc in mouse spermatozoa during maturation in the epididymis assessed by the fluorophore Zinquin.

The Zn(II)-specific fluorophore Zinquin was used to determine the regional distribution of free or loosely-bound Zn(II) in mouse spermatozoa. Spermatozoa from the testes exhibited bright fluorescence over the entire head; those from the caput epididymides generally fluoresced more brightly in the post-acrosomal region; and spermatozoa from the caudae epididymides fluoresced less brightly, with foci of fluorescence over the sperm head which were lost after extraction with Triton X-100 and hence appeared to be membrane-associated. Treatment of cauda sperm with sodium dodecyl sulfate resulted in a bright uniform Zinquin fluorescence in the heads, similar to that observed in caput sperm, indicating that the two types of sperm have similar amounts of head Zn(II) but that the availability of Zn(II) for binding Zinquin is different. By contrast, the intensity of tail fluorescence was similar in spermatozoa from different regions of the male reproductive tract and was largely unaffected by Triton X-100 extraction, consistent with an intracellular location. Similar differences were observed between caput sperm and cauda sperm in the rat. It is concluded that visualization and measurement of free or loosely-bound Zn(II) in subcellular compartments of spermatozoa should facilitate investigation of the role of this metal in the development and function of spermatozoa and abnormalities that might accompany infertility and Zn(II) deficiency.

[Pituitary adenylate cyclase activating polypeptide ameliorates the damage and inhibits the increase of intracellular calcium concentration in cultured hippocampal neurons induced by glutamate].

Hippocampal neurons cultured from 7 to 9 d in vitro were used to observe the effect of glutamate. Treatment of glutamate for 24 h greatly decreased neuronal survival and pretreatment with pituitary adenylate cyclase activating polypeptide (PACAP) significantly attenuated hippocampal neuron death induced by glutamate. Moreover, glutamate dose-dependently increased the intracellular calcium concentration in cultured hippocampal neurons, while PACAP inhibited the increase of intracellular calcium concentration induced by glutamate. PACAP 6-38, a specific PACAP type I receptor antagonist, completely inhibited the amelioration of glutamate induced death and the decrease of intracellular calcium concentration induced by PACAP in cultured hippocampal neurons. The data suggest that PACAP has a neuroprotective effect on the hippocampal neuronal damage induced by glutamate, which is related to an inhibition of glutamate-induced increase of intracellular calcium concentration and mediated by PACAP type I receptor.

[Analgesic effect and gastro-intestinal motility inhibitory action of 3-hydroxy-4-methoxy-acetophenone from Cynanchum paniculatum (Bunge) Kitagawa].

3-Hydroxy-4-methoxy-acetophenone (HMA) isolated from Cynanchum paniculatum was found to have analgesic effect and inhibitory action on the gastro-intestinal motility. The effect and action were equivalent to that of paeonol, HMA displayed a very low degree of antibacterial activity against Escherichia coli and Shigella flexneri.

Nuclear-cytoplasmic interactions during ovine oocyte maturation.

The present studies have been undertaken to investigate the interactions that occur between the nucleus and cytoplasm of ovine oocytes at various stages during meiotic maturation. We report that the nucleus of ovine fully grown dictyate stage oocytes can be efficiently removed by a microsurgical enucleation procedure. It is demonstrated that between the initiation of maturation and germinal vesicle breakdown certain newly synthesized polypeptides are selectively sequestered in the oocyte nucleus and the major sequestered polypeptide has a relative molecular mass of 28,000, which represent at least 9% of the total labelled polypeptides transferred to the oocyte nucleus during the first 4 h of maturation. The experiments provide evidence that the removal of the oocyte nucleus at various times before germinal vesicle breakdown (GVBD) does not prevent the major series of changes in protein synthesis that occurs after entry into a metaphase. We conclude therefore that the mixing of the nucleoplasm and cytoplasm is not essential for the initiation or progression of the protein reprogramming process during maturation. In addition, the experiments show that the development of the ability to condense chromatin during ovine oocyte maturation is independent of the oocyte nucleus. The combined results strongly support the hypothesis that the extensive series of translational changes that occur in oocytes during maturation are controlled by cytoplasmic rather than nuclear factors.

The effect of castration and androgen replacement on rat pituitary follicle stimulating hormone investigated by reversed-phase high performance liquid chromatography.

Rat follicle stimulating hormone (rFSH) was analysed in extracts of rat anterior pituitary using a newly developed separation method of reversed-phase high performance liquid chromatography (RP-HPLC) and post-column detection of immunoreactivity (rFSH-IR) by radioimmunoassay (RIA) or immunoprecipitation with a specific anti-rFSH antibody. Good separation of rFSH was achieved on a Waters Radial-Peak 8MBC18 cartridge with a linear (60 min) gradient from 10-60% acetonitrile at constant 0.1% trifluoroacetic acid concentration. The elution profiles of radioiodinated purified rFSH [(125I)rFSH] as well as extracts of adult male and female rat pituitaries contained two main immunoreactive components (peaks 1 and 2) which eluted at 16-17 min and 21 min respectively. In the (125I)rFSH elution profile, the peaks of immunobinding corresponded to two major peaks of radioactivity. The male pituitary showed greater binding to anti-rFSH-serum of peak 1 and 2 eluate compared to female pituitary. At day 20 after orchidectomy, the amount of immunoreactive rFSH (rFSH-IR) increased in both peaks (by 30% in peak 1 and 180% in peak 2); however, the rFSH-IR level in peak 2 appeared to be more variable compared with that in peak 1. Subcutaneous implantation of 5 mg or 15 mg of dihydrotestosterone (DHT) at the time of orchidectomy prevented the post-castration elevation of rFSH-IR at 10 and 20 days after surgery. The precise nature and physiological significance of the two elution peaks are not clear at this time; however, only peak 2 showed immunoreactivity with anti-rLH-alpha-subunit antibody (AFP 87713681) and the elution time of peak 2 coincided with that of (125I) rLH-alpha subunit (AFP-7264 B).(ABSTRACT TRUNCATED AT 250 WORDS)

Xenopus and chicken sperm contain a cytosolic soluble protein factor which can trigger calcium oscillations in mouse eggs.

There is evidence showing that at fertilization the sperm introduces into egg cytoplasm a protein-based cytosolic factor, which serves as the physiological trigger for inducing Ca(2+) oscillations in mammalian eggs. Here we show that sperm of nonmammalian vertebrates also contain a cytosolic protein factor that can induce Ca(2+) oscillations when introduced into mammalian eggs. We have observed that cytosolic extracts derived from Xenopus or chicken sperm could induce mouse eggs to undergo Ca(2+) oscillations similar to those induced by bovine sperm extracts. The factor responsible for inducing Ca(2+) oscillations was of high molecular weight and heat- or proteinase K-labile. We show that 0.5 chicken sperm-equivalents or 1-2 Xenopus sperm-equivalents of the extracts had enough activity to trigger Ca(2+) oscillations in mouse eggs. Our findings illustrate that although Xenopus, chicken, and mammals are evolutionarily divergent species, the function of the sperm protein factor in triggering Ca(2+) oscillations in mammalian eggs appears not to be species specific in vertebrates.

[Intracellular Ca2+ distribution and its role in mouse oocytes maturation].

To determine the role of calcium and calmodulin in mouse oocyte maturation, we examined the distribution of intracellular calcium during mouse oocyte maturation by using Mira Cal Imaging System. The calcium was present homogeneously in oocytes with intact germinal vesicle (GV) and accumulated around the nuclear region after GV breakdown(GVBD). The high level of calcium disappeared 6 hours later after GVBD. In the presence of 50 mumol/L BAPTA/AM, we failed to observe this phenomena. All eggs treated with 20 mumol/L W7, an antagonist of calmodulin, 50 mumol/L BAPTA/AM, a calcium chelator, could not develop to metaphase II (MII), although GVBD was not affected. We also detected the activity of a cytoplasmic maturation-promoting factor (MPF). W7 and BAPTA/AM had no effects on the rise of MPF activity in the course of maturation. We suggest that compartment distribution of calcium around nuclear region plays an important role in mouse oocyte maturation.

Flowering plant sperm contains a cytosolic soluble protein factor which can trigger calcium oscillations in mouse eggs.

There is evidence showing that the sperm-induced Ca(2+) oscillations in mammalian eggs at fertilization are triggered by a sperm-derived protein factor. It was established recently that the activity of the putative sperm protein in causing Ca(2+) oscillations in mammalian eggs is not species-specific in vertebrates (1, 16). Here we report that cytosolic soluble extracts derived from flowering plant sperms in Brassica campestris can also induce fertilization-like Ca(2+) oscillations when microinjected into mouse eggs. The factor responsible for inducing Ca(2+) oscillations in the plant sperm was sperm-specific and heat- or trypsin-labile. Eight to ten sperm equivalents of the plant sperm extracts had enough activity to trigger Ca(2+) oscillations in mouse eggs. Our study suggests that, although plant and mammal are evolutionary divergent species, the activity of the putative sperm protein factor in triggering Ca(2+) signaling in mammalian eggs is not specific to the animal kingdom.

Fine-scale mapping using Hardy-Weinberg disequilibrium.

Hardy-Weinberg disequilibrium (HWD) among affected individuals has recently been proposed for fine-scale mapping of disease susceptibility genes. We investigate the statistical properties of several available HWD measures and develop a new HWD measure J for fine-scale mapping. It is shown both theoretically and through simulations that the available HWD measures depend not only on the genetic distance between the marker locus of interest and the disease susceptibility locus, but also on the allele frequencies at the marker locus. On the contrary, the new measure is not affected by the allele frequencies at the marker locus under the following assumptions: (a) there is initial complete linkage disequilibrium between the marker and the disease loci, (b) there are no new mutations at the marker and the disease loci, and (c) the population under study is large. We develop a novel method to estimate the location of the disease susceptibility gene based on the HWD measure J. The estimator is robust to low mutation rates at the marker and the disease loci. We compare the standard error of the estimated disease gene loci using P excess for case-control studies with the standard error using J for case-only studies under various disease models. The newly developed method is successfully applied to a data set on hereditary haemochromatosis (HH).

Transmission/disequilibrium tests for quantitative traits.

The transmission/disequilibrium test (TDT) is a powerful method of locating disease genes. The TDT was originally proposed for use in studies of qualitative traits in families with both parents available. Recently, the TDT has been extended to studies of qualitative traits in sibships without parents available and in families with one parent available. It has also been extended for use in studies of quantitative traits in families with both parents available and in sibships with multiple offspring. In this paper, we first propose a new class of TDT-type tests for linkage in the presence of linkage disequilibrium for use in studies of families with both parents available. The TDT of Spielman et al. (1993) for qualitative traits and the TDT of Rabinowitz (1997) for quantitative traits are special cases of the new tests. Second, we propose a new class of TDT-type tests for linkage for use in studies of families with one parent available. Third, we study the validity and the power of the tests using simulations. Finally, we propose a method of combining data from different types of families. The combined test is valuable and allows researchers full use of the available data in detecting linkage between a marker locus and an unobservable quantitative trait locus. An important feature of the tests proposed in this paper is that no assumptions on the distribution of the quantitative traits are needed.

A comparison of intracellular changes in porcine eggs after fertilization and electroactivation.

The experiments compare intracellular changes in porcine eggs induced by electrical activation with those induced by sperm penetration. Adequate electrostimulation induces changes in both cortical granule exocytosis and protein synthesis similar to those induced by sperm during fertilization. However, ionic changes induced by electrostimulation differ markedly from those initiated at fertilization. Thus, dynamic video imaging using Fura-2 as a Ca2+ probe provides evidence that parthenogenetic activation induced by electrostimulation is initiated by a single sharp rise in the concentration of intracellular free calcium ([Ca2+]i) in the egg. The intracellular Ca2+ transient increase is triggered by an influx of extracellular Ca2+ immediately after electrostimulation. The amplitude of the intracellular Ca2+ transient increase is a function both of the extracellular Ca2+ concentration and of electric field parameters (field strength and pulse duration). Imaging demonstrates further that a single electrical pulse can only induce a single Ca2+ transient which usually lasts three to five minutes; no further Ca2+ transients are observed unless additional electrical stimuli are applied. By contrast, sperm-induced activation is characterised by a series of Ca2+ spikes which continue for at least 3 hours after sperm-egg fusion. The pattern of Ca2+ spiking after fertilization is not consistent during this period but changes both in frequency and amplitude. Overall, the results demonstrate that, although electrostimulation induces both cortical granule exocytosis and protein reprogramming in porcine eggs, it does not reproduce the pattern of [Ca2+]i changes induced by sperm entry at fertilization.

Ca(2+) oscillations induced by a cytosolic sperm protein factor are mediated by a maternal machinery that functions only once in mammalian eggs.

At fertilization in mammals, the sperm activates the egg by inducing a series of oscillations in the intracellular free Ca(2+) concentration. There is evidence showing that this oscillatory event is triggered by a sperm-derived protein factor which diffuses into egg cytoplasm after gamete membrane fusion. At present the identity of this factor and its precise mechanism of action is unknown. Here, we studied the specificity of action of the sperm factor in triggering Ca(2+) oscillations in mammalian eggs. In doing so, we examined the patterns of Ca(2+) signaling in mouse eggs, zygotes, parthenogenetic eggs and maturing oocytes following the stimulation of bovine sperm extracts which contain the sperm factor. It is observed that the sperm factor could induce Ca(2+) oscillations in metaphase eggs, maturing oocytes and parthenogenetically activated eggs but not in the zygotes. We present evidence that Ca(2+) oscillations induced by the sperm factor require a maternal machinery. This machinery functions only once in mammalian oocytes and eggs, and is inactivated by sperm-derived components but not by parthenogenetic activation. In addition, it is found that neither InsP(3) receptor sensitivity to InsP(3) nor Ca(2+) pool size are the determinants that cause the fertilized egg to lose its ability to generate sperm-factor-induced Ca(2+) oscillations at metaphase. In conclusion, our study suggests that the orderly sequence of Ca(2+) oscillations in mammalian eggs at fertilization is critically dependent upon the presence of a functional maternal machinery that determines whether the sperm-factor-induced Ca(2+) oscillations can persist.

Changes in intracellular calcium concentration in bovine oocytes following penetration by spermatozoa.

The pattern of changes in intracellular calcium concentration ([Ca2+]i) in bovine oocytes after penetration by spermatozoa was determined. Dynamic video imaging, using Fura-2 as a probe for intracellular free calcium, showed that activation of oocytes by spermatozoa induced multiple transient increases in [Ca2+]i with a spike interval of 24.2 +/- 7.3 min, and that the early transient increases were propagated throughout the oocytes in the form of a wave. Calcium transients at fertilization are involved in the induction of cortical granule exocytosis and the resultant block to polyspermy. The hypothesis that the inhibition of Ca2+ release from inositol trisphosphate-sensitive stores would inhibit exocytosis and increase polyspermy was tested by injecting oocytes before fertilization with heparin, a potent inhibitor of inositol trisphosphate-activated Ca2+ release. There was no significant difference after fertilization in either [Ca2+]i spikes or in polyspermy rates between control and experimental groups injected with low molecular mass heparin up to a final cytoplasmic concentration of 400 mumol l-1. We conclude that inositol trisphosphate-independent Ca2+ stores may be mobilized during the fertilization of bovine oocytes.

Spontaneous and fertilization-induced Ca2+ oscillations in mouse immature germinal vesicle-stage oocytes.

Mouse germinal vesicle (GV)-stage oocytes not only show Ca2+ oscillations in response to fertilization but also exhibit spontaneous Ca2+ oscillations during meiotic maturation in vitro. Spontaneous Ca2+ oscillations were entirely suppressed by microinjection of heparin (25 microM final intracellular concentration), an antagonist of inositol trisphosphate (IP3) receptor, whereas fertilization-induced Ca2+ oscillations were only partially inhibited by heparin even at a high dosage of 600 microM. Inhibition of endogenous IP3 generation by antagonizing phospholipase C using U73122 (20 microM final concentration) also failed to suppress the generation of fertilization-induced Ca2+ transients, suggesting that the two types of Ca2+ oscillations do not have the same dependence on IP3-induced Ca2+ release. In addition, spontaneous Ca2+ oscillations require the presence of intact GV whereas fertilization-induced Ca2+ oscillations are independent of the GV but require cytoplasm, since enucleation eliminated only spontaneous Ca2+ oscillations but not fertilization-induced Ca2+ oscillations. These results suggest that IP3-induced Ca2+ release is the primary mechanism responsible for spontaneous Ca2+ oscillations. Sperm-induced Ca2+ oscillations, however, may employ more complex mechanisms during fertilization.

Supervised intermittent chemotherapy for pulmonary tuberculosis in a rural area of China.

In order to ensure regularity of ambulatory treatment of new cases of pulmonary tuberculosis, a fully supervised intermittent chemotherapy regimen was tried in two rural counties of Beijing. The bare-foot doctors of the village health co-operatives were designated to administer and supervise treatment. The regimen consisted of isoniazid and streptomycin daily for 1 month, then every 3 days for 5 months and then every 5 days for a total of 12 or 18 months. For smear-negative cases the daily phase was omitted. The compliance rate among 229 patients in 1 year was 99.4%. The sputum conversion rate among 104 cases harbouring sensitive bacilli was 95.2%. Discontinuation of the regimen due to side-effects as necessary in 3 cases (1.3%). Since 1979, this treatment programme has been adopted in the whole rural area of Beijing, and the coverage rate among newly diagnosed smear-positive cases in 1983 reached 90%. A reserve regimen consisting of rifampicin and ethambutol for patients who do not convert their sputum after 6 months of treatment with isoniazid and streptomycin was added. The overall conversion rate achieved in 1981 was 97.8%. The average overall cost of drugs for each patient treated in this treatment programme was 49 yuan (RMB), about $24.00 U.S.