A comparative analysis of sivelestat sodium hydrate and ulinastatin combination therapy in the treatment of sepsis with acute respiratory distress syndrome.

OBJECTIVE: This comparative analysis aimed to investigate the efficacy of Sivelestat Sodium Hydrate (SSH) combined with Ulinastatin (UTI) in the treatment of sepsis with acute respiratory distress syndrome (ARDS). METHODS: A control group and an observation group were formed with eighty-four cases of patients with sepsis with ARDS, with 42 cases in each group. The control group was intravenously injected with UTI based on conventional treatment, and the observation group was injected with SSH based on the control group. Both groups were treated continuously for 7 days, and the treatment outcomes and efficacy of both groups were observed. The Murray Lung Injury Score (MLIS), Sequential Organ Failure Assessment (SOFA), and Acute Physiology and Chronic Health Evaluation II (APACHE II) were compared. Changes in respiratory function, inflammatory factors, and oxidative stress indicators were assessed. The occurrence of adverse drug reactions was recorded. RESULTS: The total effective rate in the observation group (95.24%) was higher than that in the control group (80.95%) (P < 0.05). The mechanical ventilation time, intensive care unit (ICU) hospitalization time, and duration of antimicrobial medication in the observation group were shorter and multiple organ dysfunction syndrome incidence was lower than those in the control group (P < 0.05). The mortality rate of patients in the observation group (35.71%) was lower than that in the control group (52.38%), but there was no statistically significant difference between the two groups (P > 0.05). MLIS, SOFA, and APACHE II scores in the observation group were lower than the control group (P < 0.05). After treatment, respiratory function, inflammation, and oxidative stress were improved in the observation group (P < 0.05). Adverse reactions were not significantly different between the two groups (P > 0.05). CONCLUSION: The combination of SSH plus UTI improves lung injury and pulmonary ventilation function, and reduces inflammation and oxidative stress in patients with sepsis and ARDS.

Multimodal ChatGPT-4V for Electrocardiogram Interpretation: Promise and Limitations.

This study evaluated the capabilities of the newly released ChatGPT-4V, a large language model with visual recognition abilities, in interpreting electrocardiogram waveforms and answering related multiple-choice questions for assisting with cardiovascular care.

Can DALL-E 3 Reliably Generate 12-Lead ECGs and Teaching Illustrations?

The recent integration of the latest image generation model DALL-E 3 into ChatGPT allows text prompts to easily generate the corresponding images, enabling multimodal output from ChatGPT. We explored the feasibility of DALL-E 3 for drawing a 12-lead ECG and found that it can draw rudimentary 12-lead electrocardiograms (ECG) displaying some of the parameters, although the details are not completely accurate. We also explored DALL-E 3's capacity to create vivid illustrations for teaching resuscitation-related medical knowledge. DALL-E 3 produced accurate CPR illustrations emphasizing proper hand placement and technique. For ECG principles, it produced creative heart-shaped waveforms tying ECGs to the heart. With further training, DALL-E 3 shows promise to expand easy-to-understand visual medical teaching materials and ECG simulations for different disease states. In conclusion, DALL-E 3 has the potential to generate realistic 12-lead ECGs and teaching schematics, but expert validation is still needed.

Nanocarbon or indocyanine green: Which is superior for gasless transaxillary endoscopic thyroidectomy to protect the parathyroid gland?

Background: Damage to the parathyroid glands remains a frequent complication after thyroidectomy, often resulting in hypoparathyroidism. Accordingly, identifying the parathyroid glands during thyroid surgical procedures is indispensable to prevent accidental surgical removal. Methods: The participants were randomly divided into three groups (indocyanine green [ICG], nanocarbon [NC], and control group). To identify and protect parathyroid glands during neck lymph node dissection in patients with thyroid cancer, IG was intravenously administered to the ICG group, whereas the NC group received an intra-thyroid injection of the NC suspension before dissection. IG was intravenously administered to each group after dissection. Subsequently, we analyzed surgical outcomes, including operative time, number of lymph nodes, serum calcium, and number of parathyroid glands. Results: We included 30 patients who underwent gasless transaxillary endoscopic thyroidectomy for thyroid cancer. Based on our findings, a greater number of parathyroid glands (P < 0.01) and higher postoperative parathyroid hormone (PTH) levels were detected in the NC and ICG groups than those in the control group (P < 0.01). The number of parathyroid glands and postoperative PTH levels in the NC group were higher than those in the ICG group (P < 0.01). Conclusions: Gasless transaxillary endoscopic thyroidectomy with NC and ICG for thyroid cancer could effectively protect the parathyroid gland and afford satisfactory clinical efficacy. NC could offer an advantage over ICG for protecting the parathyroid gland.

MicroRNA-181b blocks gensenoside Rg3-mediated tumor suppression of gallbladder carcinoma by promoting autophagy flux via CREBRF/CREB3 pathway.

BACKGROUND: Gallbladder cancer (GBC) is the seventh most common gastrointestinal cancer. Suppression of autophagy contributes to cell death of gallbladder cancer. Gensenoside Rg3 sensitizes tumor cells to chemotherapeutic agents through autophagy inhibition. However, its role mechanism on the progression of GBC remains vague. The present study is aimed to explore the functional action of Rg3 on GBC progression. METHODS: Expression of miR-181b and CREBRF in human gallbladder carcinoma specimen were determined by western blotting and qRT-PCR. Biological character of tumor cells were assessed by FACS, CCK8 and xenograft assays, respectively. Dual luciferase assay was employed to explore the targeting site of miR-181b. Autophagy flux was detected by IF staining. RESULTS: MiR-181b expression was increased, while CREBRF expression was reduced in GBC specimens compared to adjacent normal tissues. Based on Catalogue of Somatic Mutations in Cancer (COSMIC) database (408 GBC samples), there was negative correlation between hsa-miR-181b-5p/-3p and CREBRF which was a direct targeting of miR-181b. miR-181b mimic promoted cell proliferation and autophagy, restrained cell apoptosis by regulating CREBRF/CREB3 pathway. As an anti-tumor agent, gensenoside Rg3 inhibited cell proliferation and tumor growth, while promoted cell apoptosis by inhibiting autophagy. However, exogenous miR-181b blunted Rg3-evoked anti-tumor effect possibly by inhibiting CREBRF/CREB pathway. CONCLUSION: Collectively, these data indicates that miR-181b possibly mediates the pathologic progression of GBC by CREBRF/CREB3 signaling pathways and impairs anti-tumor effects of Rg3 on GBC development, which suggests that miR-181b might be an key switch in the process of Rg3-mediated tumor cytotoxicity in the progression of GBC.

Role of Vertical Transmission of Shoot Endophytes in Root-Associated Microbiome Assembly and Heavy Metal Hyperaccumulation in Sedum alfredii.

The transmission mode of shoot-associated endophytes in hyperaccumulators and their roles in root microbiome assembly and heavy metal accumulation remain unclear. Using 16S rRNA gene profiling, we investigated the vertical transmission of shoot-associated endophytes in relation to growth and Cd/Zn accumulation of Sedum alfredii ( Crassulaceae). Endophytes were transmitted from shoot cuttings to the rhizocompartment of new plants in both sterilized (γ-irradiated) and native soils. Vertical transmission was far more efficient in the sterile soil, and the transmitted endophytes have become a dominant component of the newly established root-associated microbiome. Based on 16S rRNA genes, the vertically transmitted taxa were identified as the families of Streptomycetaceae, Nocardioidaceae, Pseudonocardiaceae, and Rhizobiaceae. Abundances of Streptomycetaceae, Nocardioidaceae, and Pseudonocardiaceae were strongly correlated with increased shoot biomass and total Cd/Zn accumulation. Inoculation of S. alfredii with the synthetic bacterial community sharing the same phylogenetic relatedness with the vertically transmitted endophytes resulted in significant improvements in plant biomass, root morphology, and Cd/Zn accumulation. Our results demonstrate that successful vertical transmission of endophytes from shoots of S. alfredii to its rhizocompartments is possible, particularly in soils with attenuated microbiomes. Furthermore, the endophyte-derived microbiome plays an important role in metal hyperaccumulation.

Successive phytoextraction alters ammonia oxidation and associated microbial communities in heavy metal contaminated agricultural soils.

Phytoextraction is an attractive strategy for remediation of soils contaminated by heavy metal (HM), yet the effects of this practice on biochemical processes involved in soil nutrient cycling remain unknown. Here we investigated the impact of successive phytoextraction with a Cd/Zn co-hyperaccumulator Sedum alfredii (Crassulaceae) on potential nitrification rates (PNRs), abundance and composition of nitrifying communities and functional genes associated with nitrification using archaeal and bacterial 16S rRNA gene profiling and quantitative real-time PCR. The PNRs in rhizosphere were significantly (P < 0.05) lower than in the unplanted soils, and decreased markedly with planting time. The decrease of PNR was more paralleled by changes in numbers of copy and transcript of archaeal amoA gene than the bacterial counterpart. Phylogenetic analysis revealed that phytoextraction induced shifts in community structure of soil group 1.1b lineage-dominated ammonia-oxidizing archaea (AOA), Nitrosospira cluster 3-like ammonia-oxidizing bacteria (AOB) and Nitrospira-like nitrite-oxidizing bacteria (NOB). A strong positive correlation was observed between amoA gene transcript numbers and PNRs, whereas root exudates showed negative effect on PNR. This effect was further corroborated by incubation test with the concentrated root exudates of S. alfredii. Partial least squares path model demonstrated that PNR was predominantly controlled by number of AOA amoA gene transcripts which were strongly influenced by root exudation and HM level in soil. Our result reveals that successive phytoextraction of agricultural soil contaminated by HMs using S. alfredii could inhibit ammonia oxidation and thereby reduce nitrogen loss.

Abscisic acid-mediated modifications of radial apoplastic transport pathway play a key role in cadmium uptake in hyperaccumulator Sedum alfredii.

Abscisic acid (ABA) is a key phytohormone underlying plant resistance to toxic metals. However, regulatory effects of ABA on apoplastic transport in roots and consequences for uptake of metal ions are poorly understood. Here, we demonstrate how ABA regulates development of apoplastic barriers in roots of two ecotypes of Sedum alfredii and assess effects on cadmium (Cd) uptake. Under Cd treatment, increased endogenous ABA level was detected in roots of nonhyperaccumulating ecotype (NHE) due to up-regulated expressions of ABA biosynthesis genes (SaABA2, SaNCED), but no change was observed in hyperaccumulating ecotype (HE). Simultaneously, endodermal Casparian strips (CSs) and suberin lamellae (SL) were deposited closer to root tips of NHE compared with HE. Interestingly, the vessel-to-CSs overlap was identified as an ABA-driven anatomical trait. Results of correlation analyses and exogenous applications of ABA/Abamine indicate that ABA regulates development of both types of apoplastic barriers through promoting activities of phenylalanine ammonialyase, peroxidase, and expressions of suberin-related genes (SaCYP86A1, SaGPAT5, and SaKCS20). Using scanning ion-selected electrode technique and PTS tracer confirmed that ABA-promoted deposition of CSs and SL significantly reduced Cd entrance into root stele. Therefore, maintenance of low ABA levels in HE minimized deposition of apoplastic barriers and allowed maximization of Cd uptake via apoplastic pathway.

Antitumor effect of ginsenoside Rg3 on gallbladder cancer by inducing endoplasmic reticulum stress-mediated apoptosis in vitro and in vivo.

Recent studies have highlighted the importance of the endoplasmic reticulum (ER) in apoptotic processes. In the present study, the traditional herbal medicine ginsenoside Rg3 was used to treat gallbladder cancer in vitro and in vivo. The underlying signaling mechanisms were investigated using various molecular biology techniques, including flow cytometry, western blot analysis, ELISA and reverse transcription-quantitative polymerase chain reaction (RT-qPCR). It was indicated that Rg3 exerted pro-apoptotic activity against the gallbladder cancer cell line GBC-SD through the ER stress-mediated signaling pathway. This was demonstrated by increased expression of phosphorylation of eukaryotic translation-initiation factor 2α (eIF2α), activating transcription factor 4 (ATF4), CCAAT/enhancer-binding protein homologous protein and lipocalin 2. In addition, eIF2α and ATF4 knockdown attenuated the pro-apoptotic effect of Rg3 by inhibiting reactive oxygen species. Furthermore, the results of RT-qPCR analysis indicated that long intergenic non-protein coding RNA-p21 was significantly upregulated following Rg3 treatment. In conclusion, the results of the present study demonstrated that Rg3 inhibited tumor growth in a GBC-SD gallbladder cancer xenograft, by upregulating the ER stress-mediated signaling pathway. Therefore, ER stress activation is suggested to mediate the antitumor effect of Rg3 in gallbladder cancer activity in vitro and in vivo.

Effect of emodin on T cell subsets in NOD mice with NaI­induced experimental autoimmune thyroiditis.

Chronic lymphocytic thyroiditis (CLT), also known as Hashimoto's thyroiditis, is an autoimmune disease in which the thyroid gland is gradually destroyed. To date, only a limited number of agents can effectively suppress thyroiditis development in CLT patients. The aim of the current study was to investigate the protective effect of emodin on experimental autoimmune thyroiditis (EAT) in mice, which is considered an excellent model for CLT. NaI was used to induce the EAT model in non­obese diabetic (NOD) mice. An ELISA method was employed to detect the TgAb level (thyroid inflammation) in the serum of the EAT mice. The T cell subsets in peripheral blood and spleen were detected by flow cytometry. The histopathological study revealed that the thyroid inflammatory cell infiltration was significantly reduced by emodin compared with the model group. In addition, ELISA assays indicated that the NaI­induced serum TgAb upregulation was dramatically revered by emodin. Moreover, the level of serum IFN­Î³ and the cell populations of CD3+CD4+IL­4+, CD3+CD4+ IFN­Î³+, CD3+CD8+IL­4+, CD3+CD8+ IFN­Î³+ T cells in peripheral blood monocytes and splenic lymphocytes were significantly increased by NaI in the model group compared with in the normal group. Nevertheless, this type of increase was markedly attenuated by emodin. To conclude, the EAT model was successfully established by treating NOD mice with NaI. Emodin indicated an inhibitory effect on the autoimmune response that was significantly different in EAT compared with control mice. Furthermore, the anti­inflammatory action of emodin on EAT mice may be mediated via the inhibition of the secretion of IFN­Î³ and the cell numbers of CD3+CD4+IL­4+, CD3+CD4+ IFN­Î³+, CD3+CD8+IL­4+ and CD3+CD8+ IFN­Î³+ T cells in the peripheral blood monocytes and splenic lymphocytes. Therefore, the data may offer valuable insight on the efficacy of treatment of CLT with emodin.

Effects of elevated CO2 and endophytic bacterium on photosynthetic characteristics and cadmium accumulation in Sedum alfredii.

Elevated CO2 and use of endophytic microorganisms have been considered as efficient and novel ways to improve phytoextraction efficiency. However, the interactive effects of elevated CO2 and endophytes on hyperaccumulator is poorly understood. In this study, a hydroponics experiment was conducted to investigate the combined effect of elevated CO2 (eCO2) and inoculation with endophyte SaMR12 (ES) on the photosynthetic characteristics and cadmium (Cd) accumulation in hyperaccumulator Sedum alfredii. The results showed that eCO2 × ES interaction promoted the growth of S. alfredii, shoot and root biomass net increment were increased by 264.7 and 392.3%, respectively, as compared with plants grown in ambient CO2 (aCO2). The interaction of eCO2 and ES significantly (P < 0.05) increased chlorophyll content (53.2%), Pn (111.6%), Pnmax (59.8%), AQY (65.1%), and Lsp (28.8%), but reduced Gs, Tr, Rd, and Lcp. Increased photosynthetic efficiency was associated with higher activities of rubisco, Ca2+-ATPase, and Mg2+-ATPase, and linked with over-expression of two photosystem related genes (SaPsbS and SaLhcb2). PS II activities were significantly (P < 0.05) enhanced with Fv/Fm and Φ(II) increased by 12.3 and 13.0%, respectively, compared with plants grown in aCO2. In addition, the net uptake of Cd in the shoot and root tissue of S. alfredii grown in eCO2 × ES treatment was increased by 260.7 and 434.9%, respectively, due to increased expression of SaHMA2 and SaCAX2 Cd transporter genes. Our results suggest that eCO2 × ES can promote the growth of S. alfredii due to increased photosynthetic efficiency, and improve Cd accumulation and showed considerable potential of improving the phytoextraction ability of Cd by S. alfredii.

Effects of Cr2O3 nanoparticles on the chlorophyll fluorescence and chloroplast ultrastructure of soybean (Glycine max).

Chromic oxide nanoparticles (Cr2O3 NPs) are widely used in commercial factories and can cause serious environmental problems. However, the mechanism behind Cr2O3 NP-induced phytotoxicity remains unknown. In this study, the effects of Cr2O3 NPs on the growth, chlorophyll fluorescence, SEM-EDS analysis, and chloroplast ultrastructure of soybean (Glycine max) were investigated to evaluate its phytotoxicity. The growth of soybean treated with various Cr2O3 NP suspensions (0.01, 0.05, 0.1, and 0.5 g L-1) was significantly inhibited. Specially, shoot and root biomass decreased by 9.9 and 46.3%, respectively. Besides, the maximum quantum yield of PSII (Fv/Fm) as well as the photochemical quenching (qP) decreased by 8-22 and 30-37%, respectively, indicating that the photosynthetic system was damaged when treated with Cr2O3 NPs. Moreover, the inhibition was confirmed by the reduction of Rubisco and MDH enzyme activity (by 54.5-86.4 and 26.7-96.5%, respectively). Overall, results indicated that the damage was caused by the destruction of chloroplast thylakoid structure, which subsequently reduced the photosynthetic rate. Our research suggests that Cr2O3 NPs can be transported and cause irreversible damage to soybean plants by inhibiting the activity of electron acceptors (NADP+) and destroying ultrastructure of chloroplasts, providing insights into plant toxicity issues. Graphical abstract ᅟ.

Endophytic bacterium Buttiauxella sp. SaSR13 improves plant growth and cadmium accumulation of hyperaccumulator Sedum alfredii.

Inoculation with endophytic bacterium has been considered as a prospective application to improve the efficiency of phytoextraction. In this study, the effect of Buttiauxella sp. SaSR13 (SaSR13), a novel endophytic bacterium isolated from the root of hyperaccumulator Sedum alfredii, on plant growth and cadmium (Cd) accumulation in S. alfredii was investigated. Laser scanning confocal microscopic (LSCM) images showed that SaSR13 was mainly colonized in the root elongation and mature zones. The inoculation with SaSR13 to Cd-treated plants significantly enhanced plant growth (by 39 and 42% for shoot and root biomass, respectively), chlorophyll contents (by 38%), and Cd concentration in the shoot and root (by 32 and 22%, respectively). SaSR13 stimulated the development of roots (increased root length, surface area, and root tips number) due to an increase in the indole-3-acid (IAA) concentrations and a decrease in the concentrations of superoxide anion (O2.-) in plants grown under Cd stress. Furthermore, inoculation with SaSR13 enhanced the release of root exudates, especially malic acid and oxalic acid, which might have facilitated the uptake of Cd by S. alfredii. It is suggested that inoculation with endophytic bacterium SaSR13 is a promising bioaugmentation method to enhance the Cd phytoextraction efficiency by S. alfredii.

Oridonin enhances in vitro anticancer effects of lentinan in SMMC-7721 human hepatoma cells through apoptotic genes.

The aim of the present study was to determine the ability of oridonin to enhance the anticancer activity of lentinan (LNT) in SMMC-7721 human hepatoma cells in vitro by using various techniques, including MTT, flow cytometry, quantitative PCR and western blot assays. The results demonstrated that 20 µg/ml was a non-toxic concentration of oridonin for L02 normal liver cells and SMMC-7721 cells, while 0-200 µg/ml of LNT only had anti-proliferative effects on SMMC-7721 cells. LNT at 100 and 200 µg/ml inhibited the growth of SMMC-7721 cells by 22.8 and 60.0%, respectively, and after addition of 20 µg/ml oridonin, the inhibitory rate of 100 and 200 µg/ml LNT was increased to 47.2 and 80.7%, respectively. Oridonin (20 µg/ml) + LNT (200 µg/ml)-treated SMMC-7721 cells showed the highest apoptotic rate, which was 40.5±2.5%, which was higher than that of cells treated with LNT only. LNT raised the mRNA and protein expression of caspase-3, -8 and -9 as well as B-cell lymphoma 2 (Bcl-2)-associated X protein, p53 and p21, while reducing the expression of Bcl-2, Bcl extra large protein, epidermal growth factor (EGF) and EGF receptor expression in SMMC-7721 cells as compared to that in control cells. Treatment with 20 µg/ml oridonin and 200 µg/ml LNT increased these changes of gene expression. From the obtained results, it may be concluded that oridonin raised the in vitro anti-cancer effects of LNT in SMMC-7721 cells. Oridonin may also be used as a sensitizing agent to increase the anticancer activity of LNT in vivo.

Structural and functional variability in root-associated bacterial microbiomes of Cd/Zn hyperaccumulator Sedum alfredii.

Interactions between roots and microbes affect plant's resistance to abiotic stress. However, the structural and functional variation of root-associated microbiomes and their effects on metal accumulation in hyperaccumulators remain poorly understood. Here, we characterize the root-associated microbiota of a hyperaccumulating (HP) and a non-hyperaccumulating (NHP) genotype of Sedum alfredii by 16S ribosomal RNA gene profiling. We show that distinct microbiomes are observed in four spatially separable compartments: the bulk soil, rhizosphere, rhizoplane, and endosphere. Both the rhizosphere and rhizoplane were preferentially colonized by Proteobacteria, and the endosphere by Actinobacteria. The rhizosphere and endophytic microbiomes were dominated by the family of Sphingomonadaceae and Streptomycetaceae, respectively, which benefited for their survival and adaptation. The bacterial α-diversity decreases along the spatial gradient from the rhizosphere to the endosphere. Soil type and compartment were strongest determinants of root-associated community variation, and host genotype explained a small, but significant amount of variation. The enrichment of Bacteroidetes and depletion of Firmicutes and Planctomycetes in the HP endosphere compared with that of the NHP genotype may affect metal hyperaccumulation. Program PICRUSt predicted moderate functional differences in bacterial consortia across rhizocompartments and soil types. The functional categories involved in membrane transporters (specifically ATP-binding cassette transporters) and energy metabolism were overrepresented in endosphere of HP in comparison with NHP genotypes. Taken together, our study reveals substantial variation in structure and function of microbiomes colonizing different compartments, with the endophytic microbiota potentially playing an important role in heavy metal hyperaccumulation.

Predictive Factors of Idiopathic Thrombocytopenic Purpura and Long-term Survival in Chinese Adults Undergoing Laparoscopic Splenectomy.

The study aimed to investigate the long-term outcomes of laparoscopic splenectomy (LS) in Chinese patients with chronic idiopathic thrombocytopenic purpura (ITP). This was a retrospective analysis of 114 patients with ITP who underwent LS from 2001 to 2013. Patients were classified according to response at last contact: complete response (CR), partial response (PR), and no response (NR). Patients with CR had the highest platelet levels and patients with NR had the lowest. A correlation was observed between postoperative peak platelet count and platelet count on 2-month postoperative (r=0.829, P<0.01). In total, 27 patients showed NR to LS. Ten patients recurred within 3 years. The 140-month response rate to LS was 68%. Multivariate analysis showed that age and postoperative platelet count were independently associated with CR/PR. In conclusion, LS achieved good outcomes in Chinese patients with ITP. Age and postoperative peak platelet were independently associated with response.

Endoplasmic reticulum stress activation mediates Ginseng Rg3-induced anti-gallbladder cancer cell activity.

In the current study, we examined the potential effect of Ginsenoside Rg3 against gallbladder cancer cells, the underlying signaling mechanisms were also studied. We demonstrated that Rg3 exerted potent cytotoxic and pro-apoptotic activity against established and primary human gallbladder cancer cells. Yet it was safe to non-cancerous gallbladder epithelial cells. At the molecular level, we showed that Rg3 induced endoplasmic reticulum (ER) stress activation, the latter was evidenced by C/EBP homologous protein (CHOP) upregulation, inositol-requiring enzyme 1 (IRE1)/PKR-like endoplasmic reticulum kinase (PERK) phosphorylations, and caspase-12 activation in gallbladder cancer cells. Reversely, the ER stress inhibitor salubrinal, the caspase-12 inhibitor z-ATAD-fmk as well as CHOP shRNA knockdown significantly attenuated Rg3-induced cytotoxicity against gallbladder cancer cells. In vivo, we showed that Rg3 oral administration significantly inhibited GBC-SD gallbladder cancer xenograft growth in nude mice, its activity was, however, compromised with co-administration of the ER stress inhibitor salubrinal. Thus, we suggest that ER stress activation mediates Ginseng Rg3-induced anti-gallbladder cancer cell activity in vitro and in vivo.