Arsenite Antiporter PvACR3 Driven by Its Native Promoter Increases Leaf Arsenic Accumulation in Tobacco.

Pteris vittata is the first-reported arsenic (As) hyperaccumulator, which has been applied to phytoremediation of As-contaminated soil. PvACR3, a key arsenite (AsIII) antiporter, plays an important role in As hyperaccumulation in P. vittata. However, its functions in plants are not fully understood. In this study, the PvACR3 gene was heterologously expressed in tobacco, driven by its native promoter (ProPvACR3). After growing at 5 µM AsIII or 10 µM AsV in hydroponics for 1-5 days, PvACR3-expression enhanced the As levels in leaves by 66.4-113 and 51.8-101%, without impacting the As contents in the roots or stems. When cultivated in As-contaminated soil, PvACR3-expressed transgenic plants accumulated 47.9-85.5% greater As in the leaves than wild-type plants. In addition, PvACR3-expression increased the As resistance in transgenic tobacco, showing that enhanced leaf As levels are not detrimental to its overall As tolerance. PvACR3 was mainly expressed in tobacco leaf veins and was likely to unload AsIII from the vein xylem vessels to the mesophyll cells, thus elevating the leaf As levels. This work demonstrates that heterologously expressing PvACR3 under its native promoter specifically enhances leaf As accumulation in tobacco, which helps to reveal the As-hyperaccumulation mechanism in P. vittata and to enhance the As accumulation in plant leaves for phytoremediation.

Fatal swine acute diarrhoea syndrome caused by an HKU2-related coronavirus of bat origin.

Cross-species transmission of viruses from wildlife animal reservoirs poses a marked threat to human and animal health 1 . Bats have been recognized as one of the most important reservoirs for emerging viruses and the transmission of a coronavirus that originated in bats to humans via intermediate hosts was responsible for the high-impact emerging zoonosis, severe acute respiratory syndrome (SARS) 2-10 . Here we provide virological, epidemiological, evolutionary and experimental evidence that a novel HKU2-related bat coronavirus, swine acute diarrhoea syndrome coronavirus (SADS-CoV), is the aetiological agent that was responsible for a large-scale outbreak of fatal disease in pigs in China that has caused the death of 24,693 piglets across four farms. Notably, the outbreak began in Guangdong province in the vicinity of the origin of the SARS pandemic. Furthermore, we identified SADS-related CoVs with 96-98% sequence identity in 9.8% (58 out of 591) of anal swabs collected from bats in Guangdong province during 2013-2016, predominantly in horseshoe bats (Rhinolophus spp.) that are known reservoirs of SARS-related CoVs. We found that there were striking similarities between the SADS and SARS outbreaks in geographical, temporal, ecological and aetiological settings. This study highlights the importance of identifying coronavirus diversity and distribution in bats to mitigate future outbreaks that could threaten livestock, public health and economic growth.

Comparison of Diatrizoate and Iohexol for Patient Acceptance and Fecal-Tagging Performance in Noncathartic CT Colonography.

OBJECTIVE: The aim of this study was to compare diatrizoate and iohexol regarding patient acceptance and fecal-tagging performance in noncathartic computed tomography colonography. METHODS: This study enrolled 284 volunteers with fecal tagging by either diatrizoate or iohexol at an iodine concentration of 13.33 mg/mL and an iodine load of 24 g. Patient acceptance was rated on a 4-point scale of gastrointestinal discomfort. Two gastrointestinal radiologists jointly analyzed image quality, fecal-tagging density and homogeneity, and residual contrast agent in the small intestine. The results were compared by the generalized estimating equation method. RESULTS: Patient acceptance was comparable between the 2 groups (3.95 ± 0.22 vs 3.96 ± 0.20, P = 0.777). The diatrizoate group had less residual fluid and stool than the iohexol group ( P = 0.019, P = 0.004, respectively). There was no significant difference in colorectal distention, residual fluid, and stool tagging quality between the 2 groups (all P 's > 0.05). The mean 2-dimensional image quality score was 4.59 ± 0.68 with diatrizoate and 3.60 ± 1.14 with iohexol ( P < 0.001). The attenuation of tagged feces was 581 ± 66 HU with diatrizoate and 1038 ± 117 HU with iohexol ( P < 0.001). Residual contrast agent in the small intestine was assessed at 55.3% and 62.3% for the diatrizoate group and iohexol group, respectively ( P = 0.003). CONCLUSIONS: Compared with iohexol, diatrizoate had better image quality, proper fecal-tagging density, and more homogeneous tagging along with comparable excellent patient acceptance, and might be more suitable for fecal tagging in noncathartic computed tomography colonography.

High-sensitivity synchronous angio-lymphography based on a speckle spectrum contrast OCT.

To achieve accurate selection and synchronous imaging of blood vessels and lymph, a speckle spectrum contrast method (SSC) based on spectral-domain optical coherence tomography (SD-OCT) is proposed in this Letter. In this method, the time-lapse optical coherence tomography (OCT) intensity signal is transformed to the Fourier frequency domain. By analyzing the frequency spectrum of the time-lapse OCT intensity signal, a parameter called SSC signal, which represents the ratio of different intervals of the high frequency to the low frequency, is utilized to extract and contrast different types of the vessels in the biological tissues. In the SSC spectrum, the SSC signals of the static tissue, lymphatic vessels, and vascular vessels can be separated in three different frequency intervals, enabling differentiation and synchronous imaging of the lymphatic-vascular vessels. A mouse ear was used to demonstrate the feasibility and efficiency of this method. By using the SSC signal as the imaging parameter, the lymphatic and blood vessels of the mouse ear are differentiated and visualized simultaneously. This study shows the feasibility of the three-dimensional (3D) synchronous angio-lymphography based on the SSC method, which provides a tool to improve the understanding for disease research and treatment.

Screening inflammatory protein biomarkers on premature infants with necrotizing enterocolitis.

OBJECTIVE: This study aimed to explore potential inflammatory biomarkers for early prediction of necrotizing enterocolitis (NEC) in premature infants. METHODS: Plasma samples were collected from premature infants with NEC (n = 30), sepsis (n = 29), and controls without infection (n = 29). The 92 inflammatory-related proteins were assessed via high-throughput OLINK proteomics platform. RESULTS: There were 11 inflammatory proteins that significate differences (p < 0.05) among NEC, sepsis and control preterm infants, which include IL-8, TRAIL, IL-24, MMP-10, CCL20, CXCL1, OPG, TSLP, MCP-4, TNFSF14 and LIF. A combination of these 11 proteins could serve as differential diagnosis between NEC and control infants (AUC = 0.972), or between NEC and sepsis infants (AUC = 0.881). Furthermore, the combination of IL-8, OPG, MCP-4, IL-24, LIF and CCL20 could distinguish Stage II and III of NEC (AUC = 0.977). Further analysis showed the combination of IL-8, IL-24 and CCL20 have the best prediction value for NEC and control (AUC = 0.947), NEC and sepsis (AUC = 0.838) and different severity of NEC (AUC = 0.842). CONCLUSION: Inflammatory proteins were different expressed in premature infants with NEC compared with controls or sepsis. Combining these proteins provide a higher diagnostic potential for preterm NEC infants.

Phosphine-Mediated Morita-Baylis-Hillman-Type/Wittig Cascade: Access to E-Configured 3-Styryl- and 3-(Benzopyrrole/furan-2-yl) Quinolinones.

A phosphine-mediated, well-designed Morita-Baylis-Hillman-type/Wittig cascade for the rapid assembly of a quinolinone framework from benzaldehyde derivatives is developed for the first time. By rationally combining I2/NIS-mediated cyclization, biologically relevant 3-(benzopyrrole/furan-2-yl) quinolinones were facilely synthesized in a one-pot process by starting from 3-styryl-quinolinones bearing an o-hydroxy/amino group, significantly expanding the chemical space of this privileged skeleton. Further utility of this protocol is illustrated by successfully performing this transformation in a catalytic manner through in situ reduction of phosphine oxide by phenylsilane.

[3 + 2] vs. [4 + 1] Annulation: revisiting mechanism studies on the phosphine-catalysed domino sequence of alkynoates and activated methylenes.

The mechanism of the phosphine-catalysed domino sequence of alkynoates and activated methylenes has been computationally studied. The computational results revealed that the [3 + 2] annulation sequence could be ruled out, due to a difficult Knoevenagel condensation of aromatic aldehydes and active methylenes. The reaction proceeds through a [4 + 1] annulation pathway, which involves a phosphine-catalysed MBH-type reaction followed by a [1,5]-proton shift and dehydration to afford vinyl phosphonium intermediates as four-carbon synthons in the annulation reaction. Then 1,3-dicarbonyls act as nucleophiles to attack vinyl phosphonium intermediates, subsequently leading to a stepwise [1,3]-proton shift and an intramolecular nucleophilic attack to close the five-member ring.

Metabolomic Analysis of Microcystis aeruginosa After Exposure to the Algicide L-Lysine.

The widespread occurrence of cyanobacteria blooms damages the water ecosystem and threatens the safety of potable water and human health. Exogenous L-lysine significantly inhibits the growth of a dominant cyanobacteria Microcystis aeruginosa in freshwater. However, the molecular mechanism of how lysine inhibits the growth of M. aeruginosa is unclear. In this study, both non-target and target metabolomic analysis were performed to investigate the effects of algicide L-lysine. The results showed that 8 mg L- 1 lysine most likely disrupts the metabolism of amino acids, especially the arginine and proline metabolism. According to targeted amino acid metabolomics analysis, only 3 amino acids (L-arginine, ornithine, and citrulline), which belong to the ornithine-ammonia cycle (OAC) in arginine metabolic pathway, showed elevated levels. The intracellular concentrations of ornithine, citrulline, and arginine increased by 115%, 124%, and 19.4%, respectively. These results indicate that L-lysine may affect arginine metabolism and OAC to inhibit the growth of M. aeruginosa.

A phosphine-catalysed one-pot domino sequence to access cyclopentene-fused coumarins.

A novel phosphine-catalysed, one-pot domino approach for the annulation of 2-formylphenyl alkynoates with activated methylene compounds to construct various cyclopentene-fused dihydrocoumarins is reported. This developed strategy provides a facile and efficient approach for the synthesis of structurally complex coumarins from inexpensive and readily available alkynoates.

Expressing Phosphate Transporter PvPht2;1 Enhances P Transport to the Chloroplasts and Increases Arsenic Tolerance in Arabidopsis thaliana.

Arsenic (As) contamination in soils is of great concerns due to its toxicity to plants. As an analogue, phosphorus plays an important role in protecting plants from As toxicity. In this study, we identified a new phosphate transporter 2 (PHT2), PvPht2;1, from As-hyperaccumulator Pteris vittata and analyzed its functions in As and P transport in a yeast mutant, and model plant Arabidopsis thalian. PvPht2;1 contained 12 transmembrane domains, sharing high identity with PHT2 genes in diverse plants. Further, independent of external P or As levels, PvPht2;1 was mainly expressed in P. vittata fronds with the expression being 3-4 folds higher than that in the roots and rhizomes. Localized to the chloroplasts based on GFP-fused PvPht2;1 in model plant tobacco, PvPht2;1 functioned as a low-affinity P transporter. Under As exposure, PvPht2;1 yeast transformants showed comparable growth with the control while high-affinity P transporter PvPht1;3 transformants showed better growth, suggesting that PvPht2;1 transported P but slower than PvPht1;3 transporter. Expressing PvPht2;1 in A. thaliana increased its shoot P concentration without influencing its As accumulation. Further, the chloroplasts' P content in transgenic A. thaliana increased by 37-59% than wild-type (WT) plants. Under As exposure, the photosynthesis of PvPht2;1-expressing A. thaliana remained stable but that of WT plants decreased. The data indicate that, under As stress, expressing PvPht2;1 in A. thaliana enhanced its P transport to the chloroplasts and protected its photosynthesis. In short, highly expressed in the fronds and not impacted by As exposure, chloroplast-located PvPht2;1 may have protected As-hyperaccumulator P. vittata from As toxicity by efficiently transporting only P to its chloroplasts.

A phosphine-mediated domino sequence of salicylaldehyde with but-3-yn-2-one: rapid access to chromanone.

Chromanone is a privileged structure with a wide range of unique biological activities. A phosphine-promoted, three-component domino sequence of salicylaldehyde with but-3-yn-2-one was well designed for the construction of the chromanone skeleton under mild conditions. As a consequence, a series of novel chromanone analogues bearing an all-carbon quaternary center were facilely assembled from commercially available starting materials with moderate to good yields, which hold promising applications in pharmacological studies. Mechanistic experiments were conducted to confirm the proposed mechanism.

Expression of New Pteris vittata Phosphate Transporter PvPht1;4 Reduces Arsenic Translocation from the Roots to Shoots in Tobacco Plants.

Arsenic-hyperaccumulator Pteris vittata is efficient in As uptake, probably through phosphate transporters (Pht). Here, for the first time, we cloned a new PvPht1;4 gene from P. vittata and investigated its role in arsenate (AsV) uptake and transport in yeast and transgenic tobacco plants. On the basis of quantitative real-time polymerase chain reaction (qRT-PCR), PvPht1;4 was abundantly expressed in P. vittata fronds and roots, with its transcripts in the roots being induced by both P deficiency and As exposure. PvPht1;4 was localized to the plasma membrane, which complemented a yeast-mutant defective in P uptake and showed higher P transport affinity than PvPht1;3. Under AsV exposure, PvPht1;4 yeast transformants showed comparable tolerance as PvPht1;3, but higher As accumulation than PvPht1;2 transformants, indicating that PvPht1;4 had considerable AsV and P transport activity. However, in soil and hydroponic experiments, PvPht1;4 expressing tobacco lines accumulated 26-44 and 37-55% lower As in the shoots than wild type plants, with lower root-to-shoot As translocation. In the roots of PvPht1;4 lines, higher glutathione (GSH) contents and expression levels of GSH synthetase gene NtGSH2 were observed. In addition, the transcripts of AsIII-GSH transporter NtABCC1 in PvPht1;4 lines were upregulated. The data suggested that PvPht1;4 lines probably detoxified As by reducing AsV to AsIII, which was then complexed with GSH and stored in the root vacuoles, thereby reducing As translocation in transgenic tobacco. Given its strong AsV transport capacity, expression of PvPht1;4 provides a new molecular approach to reduce As accumulation in plant shoots.

Expressing Arsenite Antiporter PvACR3;1 in Rice (Oryza sativa L.) Decreases Inorganic Arsenic Content in Rice Grains.

Rice (Oryza sativa) is a major food crop in the world, feeding half of the world's population. However, rice is efficient in taking up toxic metalloid arsenic (As), adversely impacting human health. Among different As species, inorganic As is more toxic than organic As. Thus, it is important to decrease inorganic As in rice to reduce human exposure from the food chain. The arsenite (AsIII) antiporter gene PvACR3;1 from As-hyperaccumulator Pteris vittata decreases shoot As accumulation when heterologously expressed in plants. In this study, three homozygous transgenic lines (L2, L4, and L7) of T3 generation were obtained after transforming PvACR3;1 into rice. At 5 µM of AsIII, PvACR3;1 transgenic rice accumulated 127%-205% higher As in the roots, with lower As translocation than wild type (WT) plants. In addition, at 20 µM of AsV, the transgenic rice showed similar results, indicating that expressing PvACR3;1 increased As retention in the roots from both AsIII and AsV. Furthermore, PvACR3;1 transgenic rice plants were grown in As-contaminated soils under flooded conditions. PvACR3;1 decreased As accumulations in transgenic rice shoots by 72%-83% without impacting nutrient minerals (Mn, Zn, and Cu). In addition, not only total As in unhusked rice grain of PvACR3;1 transgenic lines were decreased by 28%-39%, but also inorganic As was 26%-46% lower. Taken together, the results showed that expressing PvACR3;1 effectively decreased both total As and inorganic As in rice grain, which is of significance to breed low-As rice for food safety and human health.

Heterologous Expression of Pteris vittata Phosphate Transporter PvPht1;3 Enhances Arsenic Translocation to and Accumulation in Tobacco Shoots.

Arsenic-hyperaccumulator Pteris vittata is efficient in As accumulation and has been used in phytoremediation of As-contaminated soils. Arsenate (AsV) is the predominant As species in aerobic soils and is taken up by plants via phosphate transporters (Pht) including P. vittata. In this work, we cloned the PvPht1;3 full length coding sequence from P. vittata and investigated its role in As accumulation by yeast and plants. PvPht1;3 complemented a yeast P uptake mutant strain and showed a stronger affinity and transport capacity to AsV than PvPht1;2. In transgenic tobacco, PvPht1;3 enhanced AsV absorption and translocation, increasing As accumulation in the shoots under both hydroponic and soil experiments. On the basis of the expression patterns via qRT-PCR, PvPht1;3 was strongly induced by P deficiency but not As exposure. To further understand its expression pattern, transgenic Arabidopsis thaliana and soybean expressing the GUS reporter gene, driven by PvPht1;3 promoter, were produced. The GUS staining showed that the reporter gene was mainly expressed in the stele cells, indicating that PvPht1;3 was expressed in stele cells and was likely involved in P/As translocation. Taken together, the data suggested that PvPht1;3 was a high-affinity AsV transporter and was probably responsible for efficient As translocation in P. vittata. Our results suggest that expressing PvPht1;3 enhances As translocation and accumulation in plants, thereby improving phytoremediation of As-contaminated soils.

Phosphate Transporter PvPht1;2 Enhances Phosphorus Accumulation and Plant Growth without Impacting Arsenic Uptake in Plants.

Phosphorus is an important macronutrient for plant growth and is acquired by plants mainly as phosphate (P). Phosphate transporters (Phts) are responsible for P and arsenate (AsV) uptake in plants including arsenic-hyperaccumulator Pteris vittata. P. vittata is efficient in AsV uptake and P utilization, but the molecular mechanism of its P uptake is largely unknown. In this study, a P. vittata Pht, PvPht1;2, was cloned and transformed into tobacco ( Nicotiana tabacum). In hydroponic experiments, all transgenic lines displayed markedly higher P content and better growth than wild type, suggesting that PvPht1;2 mediated P uptake in plants. In addition, expressing PvPht1;2 also increased the shoot/root 32P ratio by 69-92% and enhanced xylem sap P by 46-62%, indicating that PvPht1;2 also mediated P translocation in plants. Unlike many Phts permeable to AsV, PvPht1;2 showed little ability to transport AsV. In soil experiments, PvPht1;2 also significantly increased shoot biomass without elevating As accumulation in PvPht1;2 transgenic tobacco. Taken together, our results demonstrated that PvPht1;2 is a specific P transporter responsible for P acquisition and translocation in plants. We envisioned that PvPht1;2 can enhance crop P acquisition without impacting AsV uptake, thereby increasing crop production without compromising food safety.

Heterologous Expression of Pteris vittata Arsenite Antiporter PvACR3;1 Reduces Arsenic Accumulation in Plant Shoots.

Arsenic (As) is a toxic carcinogen so it is crucial to decrease As accumulation in crops to reduce its risk to human health. Arsenite (AsIII) antiporter ACR3 protein is critical for As metabolism in organisms, but it is lost in flowering plants. Here, a novel ACR3 gene from As-hyperaccumulator Pteris vittata, PvACR3;1, was cloned and expressed in Saccharomyces cerevisiae (yeast), Arabidopsis thaliana (model plant), and Nicotiana tabacum (tobacco). Yeast experiments showed that PvACR3;1 functioned as an AsIII-antiporter to mediate AsIII efflux to an external medium. At 5 µM AsIII, PvACR3;1 transgenic Arabidopsis accumulated 14-29% higher As in the roots and 55-61% lower As in the shoots compared to WT control, showing lower As translocation. Besides, transgenic tobacco under 5 µM AsIII or AsV also showed similar results, indicating that expressing PvACR3;1 gene increased As retention in plant roots. Moreover, observation of PvACR3;1-green fluorescent protein fusions in transgenic Arabidopsis showed that PvACR3;1 protein localized to the vacuolar membrane, indicating that PvACR3;1 mediated AsIII sequestration into vacuoles, consistent with increased root As. In addition, soil experiments showed ∼22% lower As in the shoots of transgenic tobacco than control. Thus, our study provides a potential strategy to limit As accumulation in plant shoots, representing the first report to decrease As translocation by sequestrating AsIII into vacuoles, shedding light on engineering low-As crops to improve food safety.

Knocking Out OsPT4 Gene Decreases Arsenate Uptake by Rice Plants and Inorganic Arsenic Accumulation in Rice Grains.

Arsenic (As) accumulation in rice grains poses health risk to humans. Plants including rice take up arsenate (AsV) by phosphate transporters. In this study, rice phosphate transporter OsPT4 (OsPht1;4) was investigated based on two independent T-DNA insertion mutants of OsPT4 (M1 and M2), which displayed stronger AsV resistance than wild types WT1 and WT2. When cultivated in medium (+P or -P) with AsV, ospt4 mutants accumulated 16-32% lower As in plants, suggesting that OsPT4 mediates AsV uptake. Analysis of the xylem sap showed that AsV concentrations in ospt4 mutants was 20-40% lower than WT controls under -P condition, indicating OsPT4 may also mediate AsV translocation. Moreover, kinetics analysis showed that ospt4 mutants had lower AsV uptake rates than the WT controls, further proving that OsPT4 functions as an AsV transporter in rice. When grown in flooded soils with As, AsV concentrations in rice grains of ospt4 mutants decreased by 50-55%. More importantly, knocking out OsPT4 in M1 and M2 reduced inorganic As accumulation in rice grains by 20-44%, significant for controlling As exposure risk from rice. Taken together, our findings revealed a critical role of OsPT4 in AsV uptake and translocation in rice. Knocking out OsPT4 effectively decreased inorganic As accumulation in rice grains, shedding light on engineering low-As rice to enhance food safety.

An aquaporin PvTIP4;1 from Pteris vittata may mediate arsenite uptake.

The fern Pteris vittata is an arsenic hyperaccumulator. The genes involved in arsenite (As(III)) transport are not yet clear. Here, we describe the isolation and characterization of a new P. vittata aquaporin gene, PvTIP4;1, which may mediate As(III) uptake. PvTIP4;1 was identified from yeast functional complement cDNA library of P. vittata. Arsenic toxicity and accumulating activities of PvTIP4;1 were analyzed in Saccharomyces cerevisiae and Arabidopsis. Subcellular localization of PvTIP4;1-GFP fusion protein in P. vittata protoplast and callus was conducted. The tissue expression of PvTIP4;1 was investigated by quantitative real-time PCR. Site-directed mutagenesis of the PvTIP4;1 aromatic/arginine (Ar/R) domain was studied. Heterologous expression in yeast demonstrates that PvTIP4;1 was able to facilitate As(III) diffusion. Transgenic Arabidopsis showed that PvTIP4;1 increases arsenic accumulation and induces arsenic sensitivity. Images and FM4-64 staining suggest that PvTIP4;1 localizes to the plasma membrane in P. vittata cells. A tissue location study shows that PvTIP4;1 transcripts are mainly expressed in roots. Site-directed mutation in yeast further proved that the cysteine at the LE1 position of PvTIP4;1 Ar/R domain is a functional site. PvTIP4;1 is a new represented tonoplast intrinsic protein (TIP) aquaporin from P. vittata and the function and location results imply that PvTIP4;1 may be involved in As(III) uptake.

Arsenic Induced Phytate Exudation, and Promoted FeAsO4 Dissolution and Plant Growth in As-Hyperaccumulator Pteris vittata.

Arsenic hyperaccumulator Pteris vittata (PV) is efficient in taking up As and nutrients from As-contaminated soils. We evaluated the mechanisms used by PV to mobilize As and Fe by examining the impacts of As and root exudates on FeAsO4 solubilization, and As and Fe uptake in four plants: As-hyperaccumulators PV and Pteris multifida (PM), nonhyperaccumulator Pteris ensiformis (PE), and angiosperm plant tomato (Solanum lycopersicum). Phytate and oxalate were dominant in fern plants (>93%), which were 50-83, 15-42, and 0-32 mg kg(-1) phytate and 10-15, 7-26, and 4-12 mg kg(-1) oxalate for PV, PM, and PE respectively, with higher As inducing greater phytate exudation and no phytate being detected in tomato exudates. PV treated with phytate+FeAsO4 had higher As and Fe contents and larger biomass than phytate or FeAsO4 treatment, which were 340 vs 20 and 130 mg kg(-1) As in the fronds and 7900 vs 1600 and 4100 mg kg(-1) Fe in the roots. We hypothesized that As-induced phytate exudation helped PV to take up Fe and As from insoluble FeAsO4 and promoted PV growth. Our study suggests that phytate exudation may be special to fern plants, which may play an important role in enhancing As and nutrient uptake by plants, thereby increasing their efficiency in phytoremediation of As-contaminated soils.

[Effect of shift rotation culture on formation and activity of encapsulated hepatocytes aggregates].

Objective: To observe the effect of uniform and shift rotation culture on the formation and activity of the alginate-chitosan (AC) microencapsulated HepLL immortalized human hepatocytes and HepG2 cells aggregates. Methods: AC microcapsulated HepG2 and HepLL cells were randomly divided into two groups. Each group was divided into 3 subgroups according to uniform and shift rotation culture.The size and number of aggregates were observed and measured under laser confocal microscopy and inverted microscope dynamically. The amount of albumin synthesis was detected by ELISA, the clearance of ammonia was detected by colorimetry, and diazepam conversion function was detected by high performance liquid chromatography (HPLC). Results: On day 6, 8, 10, 12, 14 and 16, the number and size of the aggregates, albumin synthesis, diazepam clearance and ammonium clearance increased significantly in shift rotation culture group than in uniform group (all P<0.01). The albumin synthesis, diazepam clearance, and ammonium clearance in the microencapsulated HepLL groups were significantly higher than those of HepG2 cells at any time (all P<0.01). Conclusion: Shift rotation culture can significantly promote the formation and increase the activity of AC microencapsulated HepLL and HepG2 aggregates, and HepLL cells may be more suitable for bioartificial liver than HepG2.