Biomedical subjects

Ziyi Li

Publications and source records attributed to Ziyi Li.

14 recordsLinked to original sources

PubMed2026

highSpaClone enables copy number alteration inference and tumor subclone analysis for high-resolution spatial transcriptomics.

High-resolution spatially resolved transcriptomics (SRT) offers unprecedented opportunities to investigate tumor heterogeneity but poses substantial computational and analytical challenges. Here, we present highSpaClone, a computational framework for copy number alteration (CNA) inference and tumor subclone identification from high-resolution SRT data across multiple spatial scales. By integrating spatial constraints into CNA estimation and clonal clustering, highSpaClone enables neighboring spatial locations to share information, thereby improving the robustness of genomic signals and the accuracy of subclone delineation. Across multiple Xenium and Visium HD datasets, highSpaClone revealed unique transcriptional programs, clonal evolutionary trajectories, and distinct tumor-microenvironment interactions. Furthermore, in human colorectal cancer samples, highSpaClone detected CNA events in histologically normal epithelial regions, highlighting early genomic alterations associated with field cancerization. These findings establish highSpaClone as a scalable framework for studying clonal architecture and tumor evolution.

CP: cancer biology↗

PubMed2026

Real-world pathogen spectrum, clinical actionability, and host correlates of first-time mNGS testing in hospitalized patients with hematologic diseases.

BACKGROUND: Patients with hematologic diseases are highly susceptible to infection. Conventional tests often have low sensitivity. Metagenomic next-generation sequencing (mNGS) can detect many pathogens at once, but its clinical value depends on how the results are interpreted. It is often hard to tell true infection from colonization or contamination. METHODS: We retrospectively studied hospitalized hematologic patients Only adult patients (≥18 years) who received mNGS for the first time. Detected organisms were reclassified using a clinical actionability system. We also analyzed the relationships between mNGS findings, host characteristics, and short-term outcomes. RESULTS: A total of 134 patients were included. At least one organism was detected in 87.3% of patients, but only 58.2% had highly actionable results. Bacteria were the most common findings, followed by viruses and fungi. Mixed detections were frequent. Actionable results were seen more often in respiratory specimens than in blood specimens. Viral detection was associated with immune status. Pathogen read counts were only weakly related to inflammatory markers and did not independently predict adverse outcomes. Age was the only independent risk factor for adverse outcome. CONCLUSION: mNGS had a high detection rate in hematologic patients, but not all positive findings were clinically important. Result interpretation should take specimen type and host status into account. Pathogen read counts alone were not useful for predicting short-term outcome.

Humans↗

PubMed2026

Differentiating tuberculous pleurisy from pulmonary tuberculosis using mNGS: a multicenter cohort analysis.

BACKGROUND: Tuberculous pleurisy (TBP), a major extrapulmonary form of tuberculosis, is characterized by a paucibacillary state that makes diagnosis challenging. Metagenomic next-generation sequencing (mNGS) has emerged as a promising approach for MTB detection; however, its discriminatory value between TBP and pulmonary tuberculosis (PTB) among mNGS-confirmed cases, and its integration with clinical features for differential diagnosis, remain insufficiently defined. METHODS: This multicenter retrospective cohort included hospitalized patients with MTB-positive mNGS results from January 2020 to January 2025. As only mNGS-positive cases were included, overall mNGS diagnostic sensitivity cannot be estimated. Twelve TBP patients were matched 1:2 with twenty-four PTB patients by age and sex; patients with immunosuppressive conditions were excluded prior to matching. Clinical, laboratory, mNGS, and conventional TB test data were collected. Logistic regression and ROC analyses were performed. RESULTS: Conventional tests showed limited sensitivity in TBP despite universal mNGS positivity. MTB read counts were similar between groups (median 1976.5 vs. 990.0, P = 0.920). Pleural-derived specimens predominated in TBP (41.7% vs. 4.2%, P = 0.007). CRP demonstrated the highest individual discriminatory value (AUC = 0.658, P = 0.131), though no single predictor reached significance. A combined model (cough, fever, CRP, WBC) showed modest non-significant improvement (AUC = 0.722, overall P = 0.359; sensitivity 66.7%, specificity 83.3%). Given EPV ≈ 3, all findings are exploratory only. No significant prognostic predictors were identified in TBP; a non-significant trend toward lower lymphocyte counts was observed in patients with unfavorable outcomes (0.60 vs. 1.10 ×109/L, P = 0.115). CONCLUSIONS: Among mNGS-confirmed cases, MTB read counts were comparable between TBP and PTB. No single parameter reliably distinguished the two; a combined clinical model showed modest improvement but requires prospective validation in larger cohorts. Integrating mNGS with systematic clinical evaluation remains essential for accurate TB diagnosis.

Humans↗

PubMed2025

PoweREST: Statistical power estimation for spatial transcriptomics experiments to detect differentially expressed genes between two conditions.

Recent advancements in spatial transcriptomics (ST) have significantly enhanced biological research in various domains. However, the high cost for current ST data generation techniques restricts the large-scale application of ST. Consequently, maximization of the use of available resources to achieve robust statistical power for ST data is a pressing need. One fundamental question in ST analysis is detection of differentially expressed genes (DEGs) under different conditions using ST data. Such DEG analyses are performed frequently, but their power calculations are rarely discussed in the literature. To address this gap, we developed PoweREST, a power estimation tool designed to support the power calculation for DEG detection with 10X Genomics Visium data. PoweREST enables power estimation both before any ST experiments and after preliminary data are collected, making it suitable for a wide variety of power analyses in ST studies. We also provide a user-friendly, program-free web application that allows users to interactively calculate and visualize study power along with relevant parameters.

Gene Expression Profiling↗

PubMed2024

PoweREST: Statistical Power Estimation for Spatial Transcriptomics Experiments to Detect Differentially Expressed Genes Between Two Conditions.

Recent advancements in Spatial Transcriptomics (ST) have significantly enhanced biological research in various domains. However, the high cost of current ST data generation techniques restricts its application in large-scale population studies. Consequently, there is a pressing need to maximize the use of available resources to achieve robust statistical power. One fundamental question in ST analysis is to detect differentially expressed genes (DEGs) among different conditions using ST data. Such DEG analysis is often performed but the associated power calculation is rarely discussed in the literature. To address this gap, we introduce, PoweREST (https://github.com/lanshui98/PoweREST), a power estimation tool designed to support power calculation of DEG detection with 10X Genomics Visium data. PoweREST enables power estimation both before any ST experiments or after preliminary data are collected, making it suitable for a wide variety of power analyses in ST studies. We also provide a user-friendly, program-free web application (https://lanshui.shinyapps.io/PoweREST/), allowing users to interactively calculate and visualize the study power along with relevant the parameters.

Differentially expressed genes↗

PubMed2006

Cloned ferrets produced by somatic cell nuclear transfer.

Somatic cell nuclear transfer (SCNT) offers great potential for developing better animal models of human disease. The domestic ferret (Mustela putorius furo) is an ideal animal model for influenza infections and potentially other human respiratory diseases such as cystic fibrosis, where mouse models have failed to reproduce the human disease phenotype. Here, we report the successful production of live cloned, reproductively competent, ferrets using species-specific SCNT methodologies. Critical to developing a successful SCNT protocol for the ferret was the finding that hormonal treatment, normally used for superovulation, adversely affected the developmental potential of recipient oocytes. The onset of Oct4 expression was delayed and incomplete in parthenogenetically activated oocytes collected from hormone-treated females relative to oocytes collected from females naturally mated with vasectomized males. Stimulation induced by mating and in vitro oocyte maturation produced the optimal oocyte recipient for SCNT. Although nuclear injection and cell fusion produced mid-term fetuses at equivalent rates (approximately 3-4%), only cell fusion gave rise to healthy surviving clones. Single cell fusion rates and the efficiency of SCNT were also enhanced by placing two somatic cells into the perivitelline space. These species-specific modifications facilitated the birth of live, healthy, and fertile cloned ferrets. The development of microsatellite genotyping for domestic ferrets confirmed that ferret clones were genetically derived from their respective somatic cells and unrelated to their surrogate mother. With this technology, it is now feasible to begin generating genetically defined ferrets for studying transmissible and inherited human lung diseases. Cloning of the domestic ferret may also aid in recovery and conservation of the endangered black-footed ferret and European mink.

Animals↗

PubMed2005

Factors affecting the efficiency of embryo transfer in the domestic ferret (Mustela putorius furo).

Embryo transfer (ET) to recipient females is a foundational strategy for a number of assisted reproductive technologies, including cloning by somatic cell nuclear transfer. In an attempt to develop efficient ET in domestic ferrets, factors affecting development of transferred embryo were investigated. Unilateral and bilateral transfer of zygotes or blastocysts in the oviduct or uterus was evaluated in recipient nulliparous or primiparous females. Developing fetuses were collected from recipient animals 21 days post-copulation and examined. The percentage of fetal formation was different (P<0.05) for unilateral and bilateral transfer of zygotes (71%) in nulliparous females with bilateral transfer (56%) in primiparous recipients. The percentage (90%) of fetal formation in nulliparous recipients following unilateral transfer of blastocysts was higher (P<0.05) than that observed in primiparous recipients with bilateral ET (73%). Notably, the percentage of fetal formation was higher (P<0.05) when blastocyts were transferred as compared to zygotes (90% versus 71%). Transuterine migration of embryos occurred following all unilateral transfers and also in approximately 50% of bilateral transfers with different number of embryos in each uterine horn. These data will help to facilitate the development of assisted reproductive strategies in the ferret and could lead to the use of this species for modeling human disease and for conservation of the endangered Mustelidae species such as black-footed ferret and European mink.

Animals↗

PubMed2005

Nuclear transfer of M-phase ferret fibroblasts synchronized with the microtubule inhibitor demecolcine.

The development of reconstructed embryos following nuclear transfer (NT) appears to be dependent upon a variety of factors, including cell cycle synchronization between the donor nucleus and recipient oocyte. Here we use the microtubule inhibitor, demecolcine, to synchronize ferret fibroblasts in metaphase (M-phase) in order to match their cell cycle position with that of the recipient oocyte at the time of NT. The fibroblasts were obtained from 28-day fetuses and cultured for 1-30 days prior to NT. Fibroblast cultures were treated with 0.05 microg/ml of demecolcine for 3 hr or overnight (14-16 hr) after various times in culture to determine the optimal conditions for M-phase synchronization. The percentage of G2/M-phase cells in demecolcine-treated cultures was significantly greater than that found in untreated cultures (P<0.05). Optimally synchronized M-phase fibroblasts were collected by mitotic shake-off and evaluated for their effectiveness in NT. M-phase somatic cell-derived NT embryos reconstituted by electrofusion or microinjection underwent implantation and formed fetuses at similar rates (5.4% vs. 3.4%, and 1.8% vs. 1.2%, respectively); however, no NT embryos developed to term. In summary, these data demonstrate two important points. First, demecolcine treatment effectively synchronizes ferret fibroblasts in M-phase of the cell cycle; and second, these somatic cells are capable of driving embryo development following NT. Our results should facilitate the development of cloned ferrets as an animal model for human lung disease such as influenza and cystic fibrosis.

Analysis of Variance↗

PubMed2005

Factors affecting the electrofusion of mouse and ferret oocytes with ferret somatic cells.

The domestic ferret, Mustela putorius furos, holds great promise as a genetic model for human lung disease, provided that key technologies for somatic cell nuclear transfer (SCNT) are developed. In this report, we extend our understanding of SCNT in this species by defining conditions for efficient cell fusion by electrical pulse. Two experimental systems were employed in this study. First, in vivo-matured mouse oocytes and ferret somatic cells were used to establish general parameters for fusion. One fibroblast, or cumulus cell, was agglutinated to nucleate, zona pellucida-free, mouse oocytes, and subjected to an electrical pulse. Similar electrical pulse conditions were also tested with 1 or 2 somatic cells inserted into the perivitelline space (PVS) of intact mouse oocytes. The fusion rate for a single fibroblast with a zona-free oocyte was 80.2%, significantly higher (P < 0.05) than that observed for 1, or 2, fibroblasts placed in the PVS (52.0% and 63.8%, respectively). The fusion rate (44.1%) following insertion of two cumulus cells was significantly higher (P < 0.05) than that following insertion of one cumulus cell (25.1%). Second, in vitro-matured ferret oocytes were enucleated, and one to three fibroblasts or cumulus cells were inserted into the PVS. Zona pellucida-free ferret oocytes were fragile and excluded from the study. The fusion rates with two or three fibroblasts were 71.4% and 76.8%, respectively; significantly higher (P < 0.05) than that for one fibroblast (48.6%). This cell number-dependent difference in fusion efficiency was also observed with cumulus cells. Fusion-derived (ferret-ferret) NT embryos cleaved, formed blastocysts in vitro, and underwent early-stage fetal development following embryo transfer. The rate of development was cell type-independent, in contrast to the cell type-dependent differences observed in fusion efficiency. In conclusion, fibroblasts fused more efficiently than cumulus cells and the efficiency of single cell fusions was improved when two or more cells were inserted into the PVS. These studies define conditions for efficient cell fusion with ferret oocytes and should facilitate SCNT and the development of genetically defined animal models in this species.

Animals↗

PubMed2004

Targeted correction of single-base-pair mutations with adeno-associated virus vectors under nonselective conditions.

Recombinant adeno-associated virus (rAAV) vectors possess the unique ability to introduce genetic alterations at sites of homology in genomic DNA through a mechanism thought to predominantly involve homologous recombination. We have investigated the efficiency of this approach using a mutant enhanced green fluorescent protein (eGFP) fluorescence recovery assay that facilitates detection of gene correction events in living cells under nonselective conditions. Our data demonstrate that rAAV infection can correct a mutant eGFP transgene at an efficiency of 0.1% in 293 cells, as determined by fluorescence-activated cell-sorting analysis. Gene repair was also confirmed using clonal expansion of GFP-positive cells and sequencing of the eGFP transgene. These results support previous findings demonstrating the efficacy of rAAV for gene targeting. In an effort to improve gene-targeting efficiencies, we evaluated several agents known to increase rAAV transduction (i.e., expression of an expressed gene), including genotoxic stress and proteasome inhibitors, but observed no correlation between the level of gene repair and rAAV transduction. Interestingly, however, our results demonstrated that enrichment of G(1)/S-phase cells in the target population through the addition of thymidine moderately (approximately 2-fold) increased gene correction compared to cells in other cell cycle phases, including G(0)/G1, G(1), and G(2)/M. These results suggest that the S phase of the cell cycle may more efficiently facilitate gene repair by rAAV. Transgenic mice expressing the mutant GFP were used to evaluate rAAV targeting efficiencies in primary fetal fibroblast and tibialis muscles. However, targeting efficiencies in primary mouse fetal fibroblasts were significantly lower (approximately 0.006%) than in 293 cells, and no correction was seen in tibialis muscles following rAAV infection. To evaluate the molecular structures of rAAV genomes that might be responsible for gene repair, single-cell injection studies were performed with purified viral DNA in a mutant eGFP target cell line. However, the failure of direct cytoplasm- or nucleus-injected rAAV DNA to facilitate gene repair suggests that some aspect of intracellular viral processing may be required to prime recombinant viral genomes for gene repair events.

Animals↗

PubMed2003

Progress toward generating a ferret model of cystic fibrosis by somatic cell nuclear transfer.

Mammalian cloning by nuclear transfer from somatic cells has created new opportunities to generate animal models of genetic diseases in species other than mice. Although genetic mouse models play a critical role in basic and applied research for numerous diseases, often mouse models do not adequately reproduce the human disease phenotype. Cystic fibrosis (CF) is one such disease. Targeted ablation of the cystic fibrosis transmembrane conductance regulator (CFTR) gene in mice does not adequately replicate spontaneous bacterial infections observed in the human CF lung. Hence, several laboratories are pursuing alternative animal models of CF in larger species such as the pig, sheep, rabbits, and ferrets. Our laboratory has focused on developing the ferret as a CF animal model. Over the past few years, we have investigated several experimental parameters required for gene targeting and nuclear transfer (NT) cloning in the ferret using somatic cells. In this review, we will discuss our progress and the hurdles to NT cloning and gene-targeting that accompany efforts to generate animal models of genetic diseases in species such as the ferret.

Animals↗

PubMed2003

Nuclear transfer: progress and quandaries.

Cloning mammals by nuclear transfer is a powerful technique that is quickly advancing the development of genetically defined animal models. However, the overall efficiency of nuclear transfer is still very low and several hurdles remain before the power of this technique will be fully harnessed. Among these hurdles include an incomplete understanding of biologic processes that control epigenetic reprogramming of the donor genome following nuclear transfer. Incomplete epigenetic reprogramming is considered the major cause of the developmental failure of cloned embryos and is frequently associated with the disregulation of specific genes. At present, little is known about the developmental mechanism of reconstructed embryos. Therefore, screening strategies to design nuclear transfer protocols that will mimic the epigenetic remodeling occurring in normal embryos and identifying molecular parameters that can assess the developmental potential of pre-implantation embryos are becoming increasingly important. A crucial need at present is to understand the molecular events required for efficient reprogramming of donor genomes after nuclear transfer. This knowledge will help to identify the molecular basis of developmental defects seen in cloned embryos and provide methods for circumventing such problems associated with cloning the future application of this technology.

Animals↗

PubMed2003

Developmental capacity of ferret embryos by nuclear transfer using G0/G1-phase fetal fibroblasts.

With the ultimate goal of establishing experimental protocols necessary for cloning ferrets, the present study has established parameters for the reconstruction of ferret embryos by nuclear transfer (NT) using G0/G1-phase donor fetal fibroblasts. Cumulus-oocyte complexes were harvested from superovulated ferrets and cultured in maturation medium for 24 h. Matured oocytes were then enucleated and injected with the fibroblast nuclei derived from 14-16-h serum-starved cells. Reconstructed embryos were then activated by a combination of electric pulses and chemical stimulations. Subsequently, the reconstructed and activated embryos were either cultured in vitro or transferred to pseudopregnant ferrets to evaluate their developmental capacity in vitro and in vivo. Our results demonstrated that 56.3% of reconstructed embryos (n = 187) cleaved, while 26.0% and 17.6% developed to morula and blastocyst phases in vitro, respectively. The blastocysts derived from NT embryos demonstrated normal morphology by differentially staining as compared to normal blastocysts developed in vivo following fertilization. In vivo developmental studies at 21 days posttransplantation demonstrated 8.8% of reconstructed embryos (n = 91) implanted into the uterine lining of recipients, while 3.3% formed fetuses. However, reconstructed embryos (n = 387) failed to develop to term (42 days). These results demonstrate donor nuclei of G0/G1-phase fetal fibroblast cells can be reprogrammed to support the development of reconstructed ferret embryos in vitro and in vivo; however, a significant third-trimester block occurs preventing full-term development.

Animals↗

PubMed2002

Conditions for in vitro maturation and artificial activation of ferret oocytes.

The ferret represents an attractive species for animal modeling of lung diseases because of the similarity between ferret and human lung biology and its relatively small size and short gestation time. In an effort to establish experimental protocols necessary for cloning ferrets, optimized conditions for in vitro maturation and artificial activation of ferret oocytes were examined. Cumulus-oocyte complexes were harvested from ovaries of superovulated ferrets, and in vitro maturation was evaluated in three different culture media: medium 1 (TCM-199 + 10% FBS), medium 2 (TCM-199 + 10% FBS with eCG [10 IU/ml] and hCG [5 IU/ml]), or medium 3 (TCM-199 + 10% FBS with eCG, hCG, and 17beta-estradiol [2 microg/ml]). After 24 h of maturation in vitro, the maturation rate of oocytes cultured in medium 2 (70%, n = 79) was significantly greater (P < 0.01) than those of oocytes cultured in the other two media (27%-36%, n = 67-73). At 48 h, similar maturation rates (56%-69%, n = 76-87) were observed for all three types of media. For activation experiments, oocytes cultured in medium 2 were stimulated with electrical and chemical stimuli either individually or in combination. Treatment with cycloheximide and 6-dimethylaminopurine (6-DMAP) following electrical stimulation resulted in 43% (n = 58) of the oocytes developing to the blastocyst stage. Such an activation rate represented a significant improvement over those obtainable under other tested conditions, including individual treatment with electrical pulses (10%, n = 41), cycloheximide (3%, n = 58), or 6-DMAP (5%, n = 59). Blastocysts derived from in vitro activation appeared to be normal morphologically and were composed of an appropriate number of both inner cell mass (mean +/- SEM, 10.3 +/- 1.1; n = 11) and trophectoderm (60.8 +/- 2.9, n = 11) cells. These results have begun to elucidate parameters important for animal modeling and cloning with ferrets.

Animals↗