Manufacturing, quality control, and GLP-grade preclinical study of nebulized allogenic adipose mesenchymal stromal cells-derived extracellular vesicles.

BACKGROUND: Human adipose stromal cells-derived extracellular vesicles (haMSC-EVs) have been shown to alleviate inflammation in acute lung injury (ALI) animal models. However, there are few systemic studies on clinical-grade haMSC-EVs. Our study aimed to investigate the manufacturing, quality control (QC) and preclinical safety of clinical-grade haMSC-EVs. METHODS: haMSC-EVs were isolated from the conditioned medium of human adipose MSCs incubated in 2D containers. Purification was performed by PEG precipitation and differential centrifugation. Characterizations were conducted by nanoparticle tracking analysis, transmission electron microscopy (TEM), Western blotting, nanoflow cytometry analysis, and the TNF-α inhibition ratio of macrophage [after stimulated by lipopolysaccharide (LPS)]. RNA-seq and proteomic analysis with liquid chromatography tandem mass spectrometry (LC-MS/MS) were used to inspect the lot-to-lot consistency of the EV products. Repeated toxicity was evaluated in rats after administration using trace liquid endotracheal nebulizers for 28 days, and respiratory toxicity was evaluated 24 h after the first administration. In vivo therapeutic effects were assessed in an LPS-induced ALI/ acute respiratory distress syndrome (ARDS) rat model. RESULTS: The quality criteria have been standardized. In a stability study, haMSC-EVs were found to remain stable after 6 months of storage at - 80°C, 3 months at - 20 °C, and 6 h at room temperature. The microRNA profile and proteome of haMSC-EVs demonstrated suitable lot-to-lot consistency, further suggesting the stability of the production processes. Intratracheally administered 1.5 × 108 particles/rat/day for four weeks elicited no significant toxicity in rats. In LPS-induced ALI/ARDS model rats, intratracheally administered haMSC-EVs alleviated lung injury, possibly by reducing the serum level of inflammatory factors. CONCLUSION: haMSC-EVs, as an off-shelf drug, have suitable stability and lot-to-lot consistency. Intratracheally administered haMSC-EVs demonstrated excellent safety at the tested dosages in systematic preclinical toxicity studies. Intratracheally administered haMSC-EVs improved the lung function and exerted anti-inflammatory effects on LPS-induced ALI/ARDS model rats.

[Stoichiometric Imbalance of Abandoned Grassland Under Precipitation Changes Regulate Soil Respiration].

As a key factor of global climate change, precipitation can affect soil respiration. Microorganisms are the key drivers of soil respiration, but the relationship between microbial stoichiometry and respiration in vulnerable habitat areas under different precipitation gradients is unclear. In this study, five precipitation gradients were simulated on a typical abandoned grassland in the loess hilly region. Soil respiration, nutrients, microbial biomass, and extracellular enzymes were measured, and the microbial measurement characteristics were calculated. The results showed that:①soil respiration (SR) increased significantly under rainfed treatment but decreased significantly under D50 treatment. ②Precipitation changes affected the stoichiometric imbalance, and the N:P imbalance of the active resource pool presented a u-shaped trend, whereas the C:P imbalance changed significantly only in 2019, with a trend of P50>P25>CK>D25>D50. Additionally, the stoichiometric imbalance was caused by the soil stoichiometry. In 2019, the C:P imbalance of the active resource pool showed a trend of P50>P25>CK>D25>D50, whereas the N:P imbalance of the active resource pool showed a u-shaped trend, and the stoichiometric imbalance was caused by soil stoichiometry changes. ③Soil ß-1,4-glucosidase (BG) enzyme decreased with increasing precipitation, and the sum activities of ß-1,4-N-acetylglucosaminidase (NAG) and leucine aminopeptidase (LAP) significantly decreased during two years of rainfall reduction treatment. The activity of alkaline phosphatase (ALP) significantly increased under increasing rainfall but significantly decreased under decreasing rainfall. BG:(NAG+LAP) and BG:ALP were significantly decreased under increasing precipitation conditions but significantly increased under decreasing precipitation conditions. ④The partial least squares path model (PLS-PM) showed that precipitation had an impact on soil respiration through influencing C:P stoichiometric imbalance and soil enzyme stoichiometric ratio. These results highlight the importance of stoichiometric imbalances in regulating soil respiration and may help predict how they are caused by precipitation change control carbon cycling and nutrient flow in terrestrial ecosystems.

[Effects of Short-term Nitrogen Addition on Soil Organic Carbon Components in Robinia pseudoacacia L. Plantation].

Nitrogen (N) deposition in the context of human activities continuously affects the carbon cycle of ecosystems. The effect of N deposition on soil organic carbon is related to the differential responses of different carbon fractions. To investigate the changes in soil organic carbon fraction and its influencing factors in the context of short-term N deposition, four N addition gradients:0 (CK), 1.5 (N1), 3 (N2), and 6 (N3) g·(m2·a)-1 were set up in acacia plantations based on field N addition experiments, and the soil physicochemical properties, microbial biomass, and enzyme activities were measured in June and September. The results showed that:① exogenous N input reduced soil pH, promoted the increase in soluble organic carbon content, and increased soil nitrogen effectiveness. ② Short-term N addition significantly reduced soil organic carbon content, and the response of each component of organic carbon to N addition was different. Among them, the content of easily oxidized organic carbon was significantly reduced and reached the lowest value under the N2 treatment, with 54.4% and 48.2% reduction compared with that of the control, respectively, and the content of inert organic carbon increased, although the increase was not significant. Nitrogen addition reduced the soil carbon pool activity and improved the stability of the soil carbon pool. Soil carbon pool activity reached its lowest under the N3 and N2 treatments, with a decrease of 53.3% and 52.80%, respectively, compared to that of the control. ③Random forest modeling indicated that the soil microbial biomass stoichiometry ratio, microbial biomass carbon, and AP were the key factors driving the changes in soil organic carbon activity under short-term N addition, explaining 65.96% and 66.68% of the changes in oxidizable organic carbon and inert organic carbon, respectively. Structural equation modeling validated the results of the random forest modeling, and soil microbial biomass stoichiometric ratios significantly influenced carbon pool activity. Short-term nitrogen addition changed soil microbial biomass and its stoichiometric ratio in the acacia plantation forest mainly through two pathways, i.e., increasing soil nitrogen effectiveness and promoting soil acidification and inhibiting extracellular carbon hydrolase activity, thus changing the soil carbon fraction ratio and participating in the soil organic carbon cycling process.

[Response Characteristics of Soil Organic Carbon Pool and Its Chemical Composition During Secondary Forest Succession in the Loess Plateau].

In order to explore the characteristics of the soil organic carbon(SOC)pool and its chemical composition during the succession of secondary forests in the Loess Plateau, samples of the primary stage (Populus davidiana forest), transition stage (Populus davidiana and Quercus wutaishansea mixed forest), and top stage (Quercus wutaishansea forest) of secondary forest succession in the Huanglong Mountain forest area of the Loess Plateau in Northern Shaanxi were selected as the research object. The variation characteristics of SOC content, storage, and its chemical composition at different soil depths (0-10, 10-20, 20-30, 30-50, and 50-100 cm) were analyzed. The results showed that:① the contents and storage of SOC increased significantly with the secondary forest succession process (P<0.05). The content of SOC decreased significantly with the increase in soil depth, and the storage of SOC increased from 64.8 Mg·hm-2 in the primary stage to 129.2 Mg·hm-2 in the top stage, with an increase of 99%. ② During the succession of secondary forests, in the surface (0-30 cm) soil organic carbon, the relative content of aliphatic carbon components that have a simple structure and can be decomposed more easily decreased, and the relative content of aromatic carbon components that have a complex structure and cannot be decomposed easily increased, indicating that the chemical composition of organic carbon stability of surface-layer soil increased significantly with the process of secondary forest succession. However, the stability of the chemical composition of SOC in the deep layer (30-100 cm) first increased and then decreased, that is, the transition stage>the top stage>the primary stage. ③In the process of secondary forest succession, the stability of SOC chemical composition in the primary stage and transition stage increased significantly with the increase in soil depth. The top stage tended to be stable, and the deep soil carbon stability decreased slightly. ④ Pearson correlation analysis showed that during the secondary forest succession process, SOC storage and chemical composition stability were significantly negatively correlated with soil total phosphorus content. In general, the content and storage of SOC in the 0-100 cm soil increased significantly during the secondary forest succession, playing the role of a "carbon sink." The stability of the chemical composition of SOC in the surface layer (0-30 cm) increased significantly, but in the deep layer (30-100 cm), it increased first and then decreased.

[Mineralization Characteristics of Soil Organic Carbon and Its Relationship with Organic Carbon Components in Artificial Robinia pseudoacacia Forest in Loess Hilly Region].

In order to explore the characteristics of organic carbon mineralization and the variation law of organic carbon components of an artificial forest in a loess hilly area, an artificial Robinia pseudoacacia forest restored for 13 years and the adjacent slope farmland were selected as the research objects, and indoor culture experiments under three different temperature treatments (15, 25, and 35℃) were carried out. The results indicated that the mineralization rate of soil organic carbon decreased sharply at first and then stabilized. The cumulative release of organic carbon increased rapidly in the initial stage of culture and gradually slowed in the later stage. Soil organic carbon mineralization in sloping farmland was more sensitive to temperature change, and its temperature sensitivity coefficient Q10 was 1.52, whereas that in R. pseudoacacia forest land was only 1.38. According to the fitting of the single reservoir first-order dynamic equation, the soil mineralization potential Cp of R. pseudoacacia forest land and slope farmland was between 2.02-4.32 g·kg-1 and 1.25-3.17 g·kg-1, respectively, that is, the mineralization potential of the R. pseudoacacia forest was higher. During the cultivation period, the content of various active organic carbon components decreased with time, and that in the R. pseudoacacia forest land was greater than that in the slope land. The cumulative carbon release of soil was significantly positively correlated with the contents of MBC and DOC (P<0.05), and Q10 (15-25℃) was negatively correlated with the contents of SOC, EOC, and SWC (P<0.05). These results could provide some reference for the study of soil carbon sequestration in loess hilly regions under climate change.

Nebulized exosomes derived from allogenic adipose tissue mesenchymal stromal cells in patients with severe COVID-19: a pilot study.

BACKGROUND: Existing clinical studies supported the potential efficacy of mesenchymal stromal cells as well as derived exosomes in the treatment of COVID-19. We aimed to explore the safety and efficiency of aerosol inhalation of the exosomes derived from human adipose-derived MSCs (haMSC-Exos) in patients with COVID-19. METHODS: The MEXCOVID trial is a phase 2a single-arm, open-labelled, interventional trial and patients were enrolled in Jinyintan Hospital, Wuhan, China. Eligible 7 patients were assigned to receive the daily dose of haMSCs-Exos (2.0 × 108 nano vesicles) for consecutively 5 days. The primary outcomes included the incidence of prespecified inhalation-associated events and serious adverse events. We also observed the demographic data, clinical characteristics, laboratory results including lymphocyte count, levels of D-dimer and IL-6 as well as chest imaging. RESULTS: Seven severe COVID-19 related pneumonia patients (4 males and 3 females) were enrolled and received nebulized haMSC-Exos. The median age was 57 year (interquartile range (IQR), 43 year to 70 year). The median time from onset of symptoms to hospital admission and administration of nebulized haMSC-Exos was 30 days (IQR, 15 days to 40 days) and 54 d (IQR, 34 d to 69 d), respectively. All COVID-19 patients tolerated the haMSC-Exos nebulization well, with no evidence of prespecified adverse events or clinical instability during the nebulization or during the immediate post-nebulization period. All patients presented a slight increase of serum lymphocyte counts (median as 1.61 × 109/L vs. 1.78 × 109/L). Different degrees of resolution of pulmonary lesions after aerosol inhalation of haMSC-Exos were observed among all patients, more obviously in 4 of 7 patients. CONCLUSIONS: Our trial shows that a consecutive 5 days inhalation dose of clinical grade haMSC-Exos up to a total amount of 2.0 × 109 nano vesicles was feasible and well tolerated in seven COVID-19 patients, with no evidence of prespecified adverse events, immediate clinical instability, or dose-relevant toxicity at any of the doses tested. This safety profile is seemingly followed by CT imaging improvement within 7 days. Further trials will have to confirm the long-term safety or efficacy in larger population. TRIAL REGISTRATION: MEXCOVID, NCT04276987.

[Effects of Warming and Increased Precipitation on Soil Respiration of Abandoned Grassland in the Loess-Hilly Regions].

Clarifying the changing trends and driving factors of soil respiration in fragile habitats under the background of climate change is of great significance for understanding the regional carbon cycle and the conversion of ecosystem carbon source and sink functions. This research focused on grasslands that had been naturally abandoned and restored for 12 years in the loess hilly region of northern Shaanxi, using an open top chamber (OTC) and artificially increased natural rainfall to simulate climate warming and precipitation increase and their interaction. Furthermore, we used a combination of field monitoring and indoor analysis to explore soil water content, temperature, and nutrient characteristics and the response characteristics of soil respiration rate to warming and increased precipitation and further analyzed the key factors driving changes in soil respiration. The results showed that:① warming (W) significantly increased the 5 cm soil temperature, with an average increase of 1.34℃ throughout the sampling year, whereas the increased precipitation (P50%) treatment significantly reduced the 5 cm soil temperature, reducing the average 5 cm soil temperature during the entire sampling year by 0.88℃ and increasing the soil water content (SWC) at the same time. The SWC was 13.12% and 16.45% higher than that in the control (CK), respectively. In addition, compared with that in the CK, the treatment of warming and increased precipitation (WP50%) not only increased soil temperature but also increased SWC; in general, the increase in temperature and precipitation played an antagonistic effect on the influence of soil temperature and humidity. ② P50% significantly increased the content of soil organic carbon, dissolved organic carbon, and labile organic carbon, causing changes in the soil stoichiometric ratio and the distribution characteristics of labile-recalcitrant carbon components, whereas W did not have a significant impact on organic carbon. In addition, soil total nitrogen and phosphorus and available nitrogen and phosphorus nutrients were not significantly different between treatments. ③ P50% significantly increased the Rs rate, and the effect of W on the soil respiration rate mainly depended on the seasonal precipitation and temperature. It was demonstrated that warming in winter and seasons with abundant rainfall had a significant promotion effect on the soil respiration rate. The exponential fitting of soil respiration rate and 5 cm soil temperature found that the soil respiration temperature sensitivity (Q10) was the highest under the precipitation treatment, reaching 1.68, whereas the Q10 was the lowest under the warming treatment (1.50). ④ Linear regression analysis showed that soil organic carbon, dissolved organic carbon, and labile organic carbon were all significantly positively correlated with soil respiration rate. Variation partitioning analysis showed that soil temperature, SWC, and nutrient characteristics explained 64.43% of the variation in soil respiration rate. The soil temperature and SWC were the main controlling factors of the change in soil respiration rate, with an explanation degree of 31.16%. Correlation analysis also showed that there was a significant correlation between SWC, soil temperature and respiration rate, soil organic carbon, dissolved organic carbon, labile organic carbon, C:N, and C:P. In summary, the climate prediction of abandoned grassland tending toward warm temperatures and high humidity in the loess hilly region will significantly affect the regional hydrothermal environment and nutrient characteristics, change the distribution ratio of soil labile and recalcitrant carbon, and promote regional soil carbon emissions. The analysis results showed that the key factor driving the change in soil respiration rate of abandoned grassland in the loess hilly region was soil temperature and SWC characteristics.

[Changes in Soil Microbial Carbon-Degrading Enzymes and Their Relationships with Carbon Pool Components During the Restoration Process of Robinia pseudoacacia].

To reveal the change in the characteristics of soil microbial C-degrading enzyme activities and the response to the components of C during the restoration process of Robinia pseudoacacia forests in the Loess Plateau, the components of the soil C pool, C-degrading enzyme activities, and microbial metabolic entropy of R. pseudoacacia in different restoration stages were studied, and the response relationship between C-degrading enzymes and soil C components was explored. The results showed that the microbial respiration (MR) first increased and then decreased with the restored years. We found that the microbial metabolic entropy (qCO2) decreased significantly with the restored years, but the microbial entropy (qMB) increased. Soil C-degrading enzymes increased significantly in the early-stage restoration of R. pseudoacacia; however, oxidizing enzymes (PO and PER) and cellobiohydrolase (CBH) decreased in the late stage of restoration. The soil organic C and recalcitrant organic C increased significantly with the restored years; however, there was no significant difference for the labile organic C. Correlation analysis and the partial least squares-path model (PLS-PM) showed that soil C-degrading enzymes and C components were significantly correlated with microbial respiration and entropy (qCO2 and qMB), respectively. The hydrolytic enzyme (BG+CBH) was significantly positively correlated with SOC, microbial biomass C, qMB, and recalcitrant and labile organic C. The oxidizing enzyme (PO+PER) was significantly positively correlated with the soil clay and qCO2. In addition, the recalcitrant organic C was the key driver of soil microbial metabolism affected by vegetation restoration. Overall, the ecosystem of R. pseudoacacia plantations would gradually stabilize with the increase in restored years and significantly increase the sequestration effect of soil C. These results will be helpful to understand the transformation rule and regulation mechanism of the soil C pool in vulnerable habitats and provide scientific basis for the restoration and management of vegetation in the Loess Plateau.

Plant Biomass and Soil Nutrients Mainly Explain the Variation of Soil Microbial Communities During Secondary Succession on the Loess Plateau.

Soil microorganisms play an important role in the circulation of materials and nutrients between plants and soil ecosystems, but the drivers of microbial community composition and diversity remain uncertain in different vegetation restoration patterns. We studied soil physicochemical properties (i.e., soil moisture, bulk density, pH, soil nutrients, available nutrients), plant characteristics (i.e., Shannon index [HPlant] and Richness index [SPlant], litter biomass [LB], and fine root biomass [FRB]), and microbial variables (biomass, enzyme activity, diversity, and composition of bacterial and fungal communities) in different plant succession patterns (Robinia pseudoacacia [MF], Caragana korshinskii [SF], and grassland [GL]) on the Loess Plateau. The herb communities, soil microbial biomass, and enzyme activities were strongly affected by vegetation restoration, and soil bacterial and fungal communities were significantly different from each other at the sites. Correlation analysis showed that LB and FRB were significantly positively correlated with the Chao index of soil bacteria, soil microbial biomass, enzyme activities, Proteobacteria, Zygomycota, and Cercozoa, while negatively correlated with Actinobacteria and Basidiomycota. In addition, soil water content (SW), pH, and nutrients have important effects on the bacterial and fungal diversities, as well as Acidobacteria, Proteobacteria, Actinobacteria, Nitrospirae, Zygomycota, and microbial biomass. Furthermore, plant characteristics and soil properties modulated the composition and diversity of soil microorganisms, respectively. Overall, the relative contribution of vegetation and soil to the diversity and composition of soil bacterial and fungal communities illustrated that plant characteristics and soil properties may synergistically modulate soil microbial communities, and the composition and diversity of soil bacterial and fungal communities mainly depend on plant biomass and soil nutrients.

[Stability of Soil Aggregates at Different Altitudes in Qinling Mountains and Its Coupling Relationship with Soil Enzyme Activities].

To explore changes in soil aggregate stability along an elevation gradient, and its regulating factors, soil samples were taken from the 0-10 cm surface layer at 3 different elevations on Taibai Mountain. We measured and analyzed the distribution of soil aggregates, physical and chemical properties, microbial biomass, and extracellular enzymes. The results showed that: ① the soil aggregates from the 3 elevations had mean weight diameters (MWD) of 2.17 mm, 1.83 mm, and 1.82 mm (increasing elevation), and geometric mean diameters (GMD) of 1.66 mm, 1.39 mm, and 1.32 mm, respectively. ② The change in soil aggregate stability along an elevation gradient was regulated by extracellular enzymes in the soil, in particular, the LAP in soil meso-aggregate and the BG in soil micro-aggregate. ③ Microorganisms can alleviate the N limitation at high elevations by adjusting the relative production of extracellular enzymes and altering nutrient utilization efficiency, which also changes soil aggregate stability along an elevation gradient. The results of this study have important scientific significance for soil quality evaluation and ecological environment protection in Taibai Mountain.

[Effects of Farmland Abandonment on Soil Enzymatic Activity and Enzymatic Stoichiometry in the Loess Hilly Region, China].

Clarifying the characteristic of soil enzymatic activity and stoichiometry variations as well as their influencing factors following farmland abandonment have important implications for understanding soil nutrient availability after revegetation and for illuminating the underlying mechanisms of soil nutrient cycling in ecosystems. To determine microbial nutrient limitations after farmland abandonment and to explore the driving factors of the variations in soil enzymatic activity and stoichiometry along a chronosequence of abandoned farmlands (0-, 10-, 20-, and 30-year-old) in the Loess Hilly Region, China, the potential activities of carbon (C)-, nitrogen (N)-, and phosphorus (P)-acquiring enzymes, soil physicochemical properties, and plant diversity and family composition were measured. The results showed that the activities of ß-1,4-N-acetylglucosaminidase (NAG), leucine aminopeptidase (LAP), and alkaline phosphatase (ALP) increased significantly with the increasing years of land abandonment, whereas the activity of ß-1,4-glucosidase (BG) showed the opposite change trend. Additionally, the ratios of BG:(NAG+LAP) and BG:ALP had the same variation trend with BG activity, which decreased significantly with increasing time, but the ratio of (NAG+LAP):ALP showed an increasing trend and then decreased, with the highest values observed in the 20-year sites. Moreover, the vector length of soil enzymatic stoichiometry decreased significantly as the years of land abandonment inceased, suggesting a reduced microbial C limitation after farmland abandonment. The vector angles <45°were observed at farmlands (0-year sites) and 10-year sites, whereas angles >45°were detected at 20-and 30-year sites, indicating that soil microbial communities were N-limited in the first 10 years of land abandonment and thereafter were P-limited. The redundancy analysis (RDA) reveled that soil organic C content, total N content, the C:N and C:P ratios, soil pH values, and plant diversity had significant effects on soil enzymatic activity and stoichiometry. A variation partitioning analysis (VPA) further demonstrated that edaphic and vegetation factors explained 62.0% of the total variance of soil enzymatic activity and stoichiometry. It should be noted that the interaction between vegetation characteristics and soil physicochemical properties was the major factor affecting soil enzymatic activity and stoichiometry, which explained 37.1% of the variance of the soil enzyme characteristics. Collectively, the application of P fertilizer should be considered to mitigate the deficiency of available P in the ecosystem during farmland abandonment, and these findings may provide a theoretical basis for understanding the mechanisms underlying microbe-mediated biogeochemical cycles as well as guiding soil nutrient management and the sustainable development of the ecological environment.

[Leaf nutrient resorption characteristics of Robinia pseudoacacia at different ages and their response to soil nutrient availability]. / ä¸åŒæž—é¾„åˆºæ§å¶ç‰‡å »åˆ†é‡å¸æ”¶ç‰¹å¾åŠå ¶å¯¹åœŸå£¤å »åˆ†æœ‰æ•ˆæ€§çš„å“åº”.

To reveal nutrient resorption characteristics of Robinia pseudoacacia and their driving factors in hilly and gully regions, we measured the concentration of total nitrogen and total phosphorus in leaves and the concentrations and stoichiometry of organic carbon, total nitrogen, total phospho-rus, ammonium, nitrate and available phosphorus in soils of R. pseudoacacia plantations with different stand ages. We analyzed the relationship between leaf nitrogen and phosphorus resorption efficiencies and soil nutrient characteristics. The nutrients in plants and soil changed significantly with stand ages. The total and available phosphorus concentrations were low in the soil. Nitrogen resorption efficiency first increased and then decreased with the increases of stand age, with a range of 48.2%-54.0% and a mean value of 48.5%. Phosphorus resorption efficiency increased significantly with stand age, with a range of 45.2%-49.4% and a mean value of 46.9%. Nitrogen resorption efficiency showed negative response to soil nitrogen and N:P. Phosphorus resorption efficiency was significantly positively correlated to soil N:P and negatively correlated to soil available phosphorus. Our results indicated that soil nutrient availability negatively drove nutrient resorption efficiency. The strategies of leaf nutrient resorption responded strongly to soil N:P due to the N2-fixing effect and P-limitation of R. pseudoacacia.

[Relationship Between the Composition of Soil Aggregates and the Distribution of Organic Carbon Under Long-Term Abandoned Restoration].

Soil aggregates are important carriers of soil organic carbon (SOC) accumulation, and play an important role in the evaluation of soil structure and quality. Natural recovery can promote change in soil aggregate structure and quantity via the redistribution of SOC in the aggregates. Natural restoration from farmland is an important vegetation restoration model on the Loess Plateau. The changes in soil aggregate structure and soil carbon stock after natural restoration have received extensive attention. However, little is known about the continuous study of soil changes on the abandoned grassland during the recovery process. Therefore, to understand how SOC accumulates in the process of natural recovery and quantitatively analyze the contribution of aggregates to the total soil carbon pool, we selected four abandoned grasslands of different restoration ages on the Loess Plateau, China, and studied the changes in soil structure, soil total organic carbon (TOC), soil C:N, soil aggregate distribution, soil aggregate stable index (mean weight diameter, MWD; geometric mean diameter, GMD), and aggregate-associated SOC changes as well as their correlations from 0-20 cm and 20-40 cm soil layers in abandoned grasslands. In addition, we calculated the contribution of aggregates with different sizes to soil TOC stock. The results showed that:① natural restoration increased the macroaggregate amount, MWD, and GMD, but decreased the amount of microaggregate and silt-and clay-sized fractions. There are significant differences in the distribution and stability of aggregates between different soil layers; the promotion effect of the surface was higher than that of the subsurface soils. ② In the 42 years after abandoning recovery, soil TOC stock, macaggregate-and mesaggregate-associated SOC stock increased significantly, and varied with soil depth and years of abandonment (1.92 times, 10.2 times, and 3.61 times). In contrast, micaggregate-associated SOC stock decreased significantly, and silt-and clay-sized fractions-associated SOC stock showed no distinct change. In addition, natural restoration promoted the ratio of C:N; nevertheless, the ratio of C:N under the surface showed a reduced phenomenon after 42 years of abandonment. ③ The improvement in soil TOC stock depends primarily on changes in the macaggregate-associated organic carbon stocks, which account for 80% of macaggregate, and the significant increase in the amount of macaggregate is the main reason for the high contribution.The results of our study suggest that natural restoration is conducive to the accumulation of soil organic carbon, and improvement in soil structure and stability. Macroaggregate is the key factor in soil organic carbon accumulation and soil structure improvement in the process of natural restoration.

[Responses mechanism of C:N:P stoichiometry of soil microbial biomass and soil enzymes to climate change.] / åœŸå£¤å¾®ç”Ÿç‰©ç”Ÿç‰©é‡ç¢³æ°®ç£·ä¸ŽåœŸå£¤é ¶åŒ–å­¦è®¡é‡å¯¹æ°”å€™å˜åŒ–çš„å“åº”æœºåˆ¶.

Microorganisms and soil enzymes are important drivers for biogeochemical cycles in terrestrial ecosystems. Understanding the role of microorganisms in the regulation of ecosystems and the response mechanisms of microbial biomass and soil enzymes to climate change are important topic in ecology. From the perspective of climatic factors, this review introduced the roles of microorganisms and soil enzymes in the carbon, nitrogen and phosphorus cycles of terrestrial ecosystems based on the theory of ecological stoichiometry. Moreover, we synthesized the responses mechanisms of soil microbial and soil enzyme stoichiometry, i.e., changes of microbial metabolic rate, enzymatic acti-vity, microbial community structure, ecological stoichiometry of soil microbial biomass and soil enzymes, and nutrient use efficiency. Finally, we analyzed the current research inadequacies and proposed the scientific problems in this field, i.e., to comprehensively elucidate the response mecha-nism of soil microbes and soil enzymes to climate change; to examine the nutrient coupling mechanism of soil microbes and extracellular enzymes; and to explore the adaptive strategies of C:N:P stoichiometry of soil microbial biomass and soil enzymes to climate change.