Isolation and Identification of Cancer Stem Cells from Human Osteosarcom by Serum-free Three-dimensional Culture Combined with Anticancer Drugs

The cancer stem cells(CSCs)from human osteosarcoma by serum-free three-dimensional culture combined with anticancer drugs were isolated and identified.The primary cells derived from human osteosarcoma were digested by trypsin to prepare a single-cell suspension,and mixed homogeneously into 1.2% alginate gel.Single-cell alginate gel was cultured with serum-free DMEM/F12 medium.Epirubicin(0.8μg/mL)was added to the medium to enrich CSCs.After cultured conventionally for 7 to 10 days,most of cells suspended in ...

Optimization of Three-Dimensional Culture Conditions of HepG2 Cells with Response Surface Methodology Based on the VitroGel System

Objective This study optimizes three-dimensional(3D) culture conditions of HepG2 using response surface methodology(RSM) based on the VitroGel system to facilitate the cell model in vitro for liver tissues.Method HepG2 cell was 3D cultured on the VitroGel system.Cell viability was detected using Cell Counting Kit-8(CCK-8) assay of HepG2 lived cell numbers.The proliferation of HepG2 cell and clustering performance was measured via fluorescence staining test.Albumin concentration in the culture medium supernatant as an index of HepG2 cell biological function was measured with ELISA kit.Independent factor tests were conducted with three key factors:inoculated cell concentration,cultured time,and dilution degree of the hydrogel.The preliminary results of independent factor tests were used to determine the levels of factors for RSM.Result The selected optimal culture conditions are as follows:concentration of inoculated cells was4.44 × 10^(5)/mL,culture time was 4.86 days,and hydrogel dilution degree was 1:2.23.The result shows that under optimal conditions,the predicted optical density(OD) value of cell viability was 3.10 and measured 2.978 with a relative error of 3.94%.Conclusion This study serves as a reference for the 3D HepG2 culture and constructs liver tissues in vitro.Additionally,it provides the foundation for repeated dose high-throughput toxicity studies and other scientific research work.

Simplified three-dimensional culture system for long-term expansion of embryonic stem cells

AIM: To devise a simplified and efficient method for long-term culture and maintenance of embryonic stem cells requiring less frequent passaging. METHODS: Mouse embryonic stem cells(ESCs) labeled with enhanced yellow fluorescent protein were cultured in three-dimensional(3-D) self-assembling scaffolds and compared with traditional two-dimentional(2-D) culture techniques requiring mouse embryonic fibroblast feeder layers or leukemia inhibitory factor. 3-D scaffolds encapsulating ESCs were prepared by mixing ESCs with polyethylene glycol tetra-acrylate(PEG-4-Acr) and thiolfunctionalized dextran(Dex-SH). Distribution of ESCs in 3-D was monitored by confocal microscopy. Viability and proliferation of encapsulated cells during long-term culture were determined by propidium iodide as well as direct cell counts and PrestoB lue(PB) assays. Genetic expression of pluripotency markers(Oct4, Nanog, Klf4, and Sox2) in ESCs grown under 2-D and 3-D cultureconditions was examined by quantitative real-time polymerase chain reaction. Protein expression of selected stemness markers was determined by two different methods, immunofluorescence staining(Oct4 and Nanog) and western blot analysis(Oct4, Nanog, and Klf4). Pluripotency of 3-D scaffold grown ESCs was analyzed by in vivo teratoma assay and in vitro differentiation via embryoid bodies into cells of all three germ layers. RESULTS: Self-assembling scaffolds encapsulating ESCs for 3-D culture without the loss of cell viability were prepared by mixing PEG-4-Acr and Dex-SH(1:1 v/v) to a final concentration of 5%(w/v). Scaffold integrity was dependent on the degree of thiol substitution of Dex-SH and cell concentration. Scaffolds prepared using Dex-SH with 7.5% and 33% thiol substitution and incubated in culture medium maintained their integrity for 11 and 13 d without cells and 22 ± 5 d and 37 ± 5 d with cells, respectively. ESCs formed compact colonies, which progressively increased in size over time due to cell proliferation as determined by confocal microscopy and PB staining. 3-D scaffold cultured ESCs expressed significantly higher levels(P < 0.01) of Oct4, Nanog, and Kl4, showing a 2.8, 3.0 and 1.8 fold increase, respectively, in comparison to 2-D grown cells. A similar increase in the protein expression levels of Oct4, Nanog, and Klf4 was observed in 3-D grown ESCs. However, when 3-D cultured ESCs were subsequently passaged in 2-D culture conditions, the level of these pluripotent markers was reduced to normal levels. 3-D grown ESCs produced teratomas and yielded cells of all three germ layers, expressing brachyury(mesoderm), NCAM(ectoderm), and GATA4(endoderm) markers. Furthermore, these cells differentiated into osteogenic, chondrogenic, myogenic, and neural lineages expressing Col1, Col2, Myog, and Nestin, respectively. CONCLUSION: This novel 3-D culture system demonstrated long-term maintenance of mouse ESCs without the routine passaging and manipulation necessary for traditional 2-D cell propagation.

Three-dimensional Culture of Human Airway Epithelium in Matrigel for Evaluation of Human Rhinovirus C and Bocavirus Infections

Objective Newly identified human rhinovirus C （HRV-C） and human bocavirus （HBoV） cannot propagate in vitro in traditional cell culture models; thus obtaining knowledge about these viruses and developing related vaccines are difficult. Therefore, it is necessary to develop a novel platform for the propagation of these types of viruses.Methods A platform for culturing human airway epithelia in a three-dimensional （3D） pattern using Matrigel as scaffold was developed. The features of 3D culture were identified by immunochemical staining and transmission electron microscopy. Nucleic acid levels of HRV-C and HBoV in 3D cells at designated time points were quantitated by real-time polymerase chain reaction {PCR）. Levels of cytokines, whose secretion was induced by the viruses, were measured by ELISA.Results Properties of bronchial-like tissues, such as the expression of biomarkers CK5, ZO-2, and PCK, and the development of cilium-like protuberances indicative of the human respiration tract, were observed in 3D-cultured human airway epithelial （HAE） cultures, but not in monolayer-cultured cells. Nucleic acid levels of HRV-C and HBoV and levels of virus-induced cytokines were also measured using the 3D culture system.Conclusion Our data provide a preliminary indication that the 3D culture model of primary epithelia using a Matrigel scaffold in vitro can be used to propagate HRV-C and HBoV.

Irradiation Response of Adipose-derived Stem Cells under Three-dimensional Culture Condition

Objective Adipose tissue distributes widely in human body. The irradiation response of the adipose cells in vivo remains to be investigated. In this study we investigated irradiation response of adipose-derived stem cells (ASCs) under three-dimensional culture condition. Methods ASCs were isolated and cultured in low attachment dishes to form three-dimensional (3D) spheres in vitro. The neuronal differentiation potential and stem-liked characteristics was monitored by using immunofluoresence staining and flow cytometry in monolayer and 3D culture. To investigate the irradiation sensitivity of 3D sphere culture, the fraction of colony survival and micronucleus were detected in monolayer and 3D culture. Soft agar assays were performed for measuring malignant transformation for the irradiated monolayer and 3D culture. Results The 3D cultured ASCs had higher differentiation potential and an higher stem-like cell percentage. The 3D cultures were more radioresistant after either high linear energy transfer (LET) carbon ion beam or low LET X-ray irradiation compared with the monolayer cell. The ASCs’ potential of cellular transformation was lower after irradiation by soft agar assay. Conclusion These findings suggest that adipose tissue cell are relatively genomic stable and resistant to genotoxic stress.

Inhibition of mammalian target of rapamycin induces phenotypic reversion in three-dimensional cultures of malignant breast epithelial cells

Inhibition of mammalian target of rapamycin (m- TOR) is a potential method for cancer treatment. Effects of rapamycin (RAP) on the reversion of malignant breast epithelial cells were investigated on three-dimensional (3D) basement membrane extract (BME) cultures. Through continuous exposure to 20 nM of RAP, cell colony size was significantly reduced in 3D BME cultures of malignant breast epithelial cells, while normal cell colony size appeared unaffected. In unfixed 3D BME cultures of normal and RAP-treated malignant breast epithelial cells, the presence of luminal cell death was confirmed by ethidium bromide and propidium iodide labeling. Increased structural organization was observed by im- munofluorescence staining of F-actin and β-catenin in RAP-treated malignant breast epithelial cells. In monolayer cultures of normal and malignant breast epithelial cells, continuous exposure to 20 nM of RAP increased caspase 3/7 activity and decreased proliferation. Reverse transcriptase polymerase ch- ain reaction (RT-PCR) array analysis indicated a fold increase in the expression of a number of proteins related to polarity, cell-cell adhesion, and cell-matrix adhesion in the presence of RAP. Our data showed that phenotypic reversion of malignancy can be ach- ieved through RAP exposure on 3D BME cultures. This 3D BME culture system will provide correct microenvironments for observing the effects of other mTOR inhibitors on phenotypic reversion of malignant breast epithelial cells.

Three-dimensional culture system enhance the differentiation of mesenchymal stem cells into neural cells in vitro

Nerve injury repair has been an intractable issue wor.ldwide due to the non-renew able property of neurons in vivo.Cell therapy adopting me senchymal stem cells(MSC)induced neurons stands for a promising substitute treatment,which is hindered by the low diffe rentiation efficiency of current approaches.To improve the efficiency of MSC differentiation in vitro,we constructed a three-dimensional(3D)culture system by seeding human bone marrow-de rived MSC(hBMSCs)into collagen-nano fiber scaffolds which are a mixed hydrogel of collagen 1 and silk nanofiber solution and testified the diffe rentiation efficiency from MSC into neurons by chemical or cocktail induction methods.Results indicated MSCs cultured in 3D system showed good viability with neuron-like morphology in both groups.

Effect of deforolimus and VEGF on angiogenesis in endometrial stromal cells following three-dimensional culture

The presence of endometrial tissue outside of the uterine cavity is named endometriosis and is the most common gynecologic disorder in women. Determining the inhibitory effect of a Deforolimus on angiogenesis in a three-dimensional (3-D) culture of human endometrial stromal cells (hEnCs) in vitro. The important mechanism in the pathogenesis of endometriosis is angiogenesis, and deforolimus has been shown to have anti-angiogenic activity. This was an in vitro study of human endometrial stromal cells in 3-D culture of fibrin matrix. Endometrial stromal cells isolated and placed in a 3-D fibrin matrix culture system for angiogenesis with VEGF and inhibit angiogenesis by deforolimus. Finally these cells analyzed by CD31 antibodies. After 3 weeks, in cells treated with VEGF, endothelial cell branching was observed and rudimentary capillary-like structures formed. In the presence of 5μM of deforolimus, angiogenesis was reduced. The deforolimus were shown to be effective in inhibiting the mechanisms of angiogenesis.

Chinese medicinal compound delisheng has satisfactory anti-tumor activity,and is associated with up-regulation of endostatin in human hepatocellular carcinoma cell line HepG2 in three-dimensional culture

AIM: To investigate the multicellular resistance of human hepatocellular carcinoma HepG2 cells in three-dimensional culture to delisheng,5-fluorouracil and adriamycin,and the possible molecular mechanisms of delisheng. METHODS: Human hepatocellular carcinoma HepG2 cells were cultured with a liquid overlay technique. After the formation of multicellular spheroids,morphology was analyzed by phase contrast microscopy,scanning electron microscopy and transmission electron microscopy. Sensitivity of HepG2 cells to delisheng,5-fluorouracil and adriamycin was investigated by MTT assay in multicelluar spheroids and monolayers. Vascular endothelial growth factor (VEGF) and endostatin expression were analyzed in multicellular spheroids treated with delisheng,5-fluorouracil,adriamycin and negative control PBS,with immunohistochemical staining. RESULTS: Multicellular spheroids exhibited structural characteristics somewhat different to those in monolayers. The cells in three-dimensional cell culture turned out to be less sensitive to delisheng,5-fluorouracil and adriamycin than the cells cultured in monolayer. This showed that delisheng had a satisfactory cells inhibition ratio compared to 5-fluorouracil and adriamycin. Immunohistochemical staining showed that VEGF and endostatin expression was positive during growth as multicellular spheroids,and endostatin expression in spheroids with treatment of delisheng was higher than that with 5-fluorouracil,adriamycin and PBS (139.35 ± 7.83,159.23 ± 10.34,162.83 ± 3.47 and 148.48 ± 11.06,P < 0.05).CONCLUSION: Chinese medicine compound delisheng has satisfactory anti-tumor activity in HepG2 cells in three-dimensional culture,and the effects are associated with up-regulation of endostatin.

Hydrogel-supported poly(L-lactic acid) and polystyrene microsphere-based three-dimensional culture systems for in vitro cell expansion

The in vitro expansion of stem cells is important for their application in different life science fields such as cellular tissue and organ repair.An objective of this paper was to achieve static cell culture in vitro through peptide hydrogel-supported microspheres(MSs).The peptides,with their gel-forming properties,microstructures,and mechanical strengths characterized,were found to have good support for the MSs and to be injectable.The internal structures of poly(L-lactic acid)microspheres(PLLA-MSs)and polystyrene microspheres(PS-MSs)made in thelaboratory were observed and statistically analyzed in terms of particle size and pore size,following which the co-cultured MSs with cells were found to have good cell adhesion.In addition,three-dimensional(3D)culturing of cells was performed on the peptide and microcarrier composite scaffolds to measure cell viability and cell proliferation.The results showed that the peptides could be stimulated by the culture medium to self-assembly form a 3D fiber network structure.Under the peptide-Ms composite scaffold-based cell culture system,further enhancement of the cell culture effect was measured.The peptide-Ms composite scaffolds have great potential for the application in 3D cell culture and in vitro cellexpansion.

Application of three-dimensional culture systems to study mammalian spermatogenesis, with an emphasis on the rhesus monkey （Macaca mulatta）

In vitro culture of spermatogonial stem cells （SSCs） has generally been performed using two-dimensional （2D） culture systems; however, such cultures have not led to the development of complete spermatogenesis. It seems that 2D systems do not replicate optimal conditions of the seminiferous tubules （including those generated by the SSC niche） and necessary for spermatogenesis. Recently, one of our laboratories has been able to induce proliferation and differentiation of mouse testicular germ cells to meiotic and postmeiotic stages including generation of sperm in a 3D soft agar culture system （SACS） and a 3D methylcellulose culture system （MCS）. It was suggested that SACS and MCS form a special 3D microenvironment that mimics germ cell niche formation in the seminiferous tubules, and thus permits mouse spermatogenesis in vitro. In this review, we （1） provide a brief overview of the differences in spermatogenesis in rodents and primates, （2） summarize data related to attempts to generate sperm in vitro, （3） report for the first time formation of colonies/clusters of cells and differentiation of meiotic （expression of CREM-1） and postmeiotic （expression of acrosin） germ cells from undifferentiated spermatogonia isolated from the testis of prepubertal rhesus monkeys and cultured in SACS and MCS, and （4） indicate research needed to optimize 3D systems for in vitro primate spermatogenesis and for possible future application to man.

Assessment of pancreatic carcinoma cell chemosensitivity using a three-dimensional culture system

Background Monolayer cell culture models are the traditional culture models used for in vitro research of pancreatic carcinoma chemosensitivity. However, these models neglect the interactions between tumor cells and the impact of the tumor microenvironment. Such tumor cell monolayers poorly mimic the solid tumor microenvironment. The present study aimed to investigate the chemosensitivity characteristics of pancreatic cancer cells in a three-dimensional culture system by analyzing the differences in drug sensitivity between a scattered cell culture model and a multicellular spheroid culture model. Methods Three pancreatic cancer cell lines （SW1990, ASPC-1 and PCT-3） were cultured in three-dimensional collagen gels as well as in traditional two-dimensional monolayers. The chemosensitivities of the pancreatic carcinoma cells to 5-fluorouracil （5-FU）, gemcitabine, and oxaliplatin in vitro were detected by both the Cell Counting Kit-8 test and the collagen gel droplet-embedded culture drug-sensitivity test. Results In the two-dimensional culture model, differences in the chemosensitivities of the cloned pancreatic carcinoma cells and scattered cells existed for some concentrations of 5-FU, gemcitabine and oxaliplatin. In the three-dimensional culture model, there were significant differences in the chemosensitivities of the pancreatic cancer cells between the scattered cells and multicellular spheroids （P 〈0.05）. Conclusion Pancreatic carcinoma cells exhibit multicellular resistance in three-dimensional cultures.

Three-dimensional cultures of human endometrial cells on Matrigel mimic in vivo morphology

Background The regulation of endometrial physiology and morphogenesis by the paracrine effectors has been well established using in vivo studies. A more complete understanding of the endometrial function has been delayed due, in part, to a lack of appropriate culture models. In this study, we aimed to simulate the in vivo three-dimensional （3-D） growth pattern of endometrial cells using a 3-D in vitro culture system.

Anticancer Activity of Rice Callus Suspension Cultures from Aromatic Varieties and Metabolites Regulated in Treated Cancer Cell Lines

Tissue culture techniques were used to produce large amounts of bioactive compounds with medicinal potential, overcoming space and time constraints for cancer prevention. Rice callus suspension cultures(RCSC) and seed extracts prepared from aromatic rice varieties were used to evaluate the cytotoxic impact on human colon and lung cancer cell lines, as well as a normal control cell line, using Taxol as a positive control. RCSC and seed extracts from two Indian aromatic rice varieties were applied at different concentrations to treat the cancer cell lines and normal lung fibroblasts over varying time intervals. Apoptosis was assessed in 1:5 dilutions of the A549 and HT-29 cell lines treated with RCSC for 72 h, using propidium iodide staining and flow cytometry. RCSC showed a more potent cytotoxic effect than seed extracts with minimal effect on the normal cell line, in contrast to Taxol. Confocal microscopy and flow cytometry further confirmed the apoptotic effect of RCSC. Gas chromatography-mass spectrometry-based metabolic profiling identified metabolites involved in cytotoxicity and highlighted altered pathways. RCSC is proposed as an alternative source for the development of novel anticancer drugs with reduced side effects.

Oxygen tension modulates cell function in an in vitro three-dimensional glioblastoma tumor model

Hypoxia is a typical feature of the tumor microenvironment,one of the most critical factors affecting cell behavior and tumor progression.However,the lack of tumor models able to precisely emulate natural brain tumor tissue has impeded the study of the effects of hypoxia on the progression and growth of tumor cells.This study reports a three-dimensional(3D)brain tumor model obtained by encapsulating U87MG(U87)cells in a hydrogel containing type I collagen.It also documents the effect of various oxygen concentrations(1%,7%,and 21%)in the culture environment on U87 cell morphology,proliferation,viability,cell cycle,apoptosis rate,and migration.Finally,it compares two-dimensional(2D)and 3D cultures.For comparison purposes,cells cultured in flat culture dishes were used as the control(2D model).Cells cultured in the 3D model proliferated more slowly but had a higher apoptosis rate and proportion of cells in the resting phase(G0 phase)/gap I phase(G1 phase)than those cultured in the 2D model.Besides,the two models yielded significantly different cell morphologies.Finally,hypoxia(e.g.,1%O2)affected cell morphology,slowed cell growth,reduced cell viability,and increased the apoptosis rate in the 3D model.These results indicate that the constructed 3D model is effective for investigating the effects of biological and chemical factors on cell morphology and function,and can be more representative of the tumor microenvironment than 2D culture systems.The developed 3D glioblastoma tumor model is equally applicable to other studies in pharmacology and pathology.

Development of a toroidal soft x-ray imaging system and application for investigating three-dimensional plasma on J-TEXT

A toroidal soft x-ray imaging(T-SXRI)system has been developed to investigate threedimensional(3D)plasma physics on J-TEXT.This T-SXRI system consists of three sets of SXR arrays.Two sets are newly developed and located on the vacuum chamber wall at toroidal positionsφof 126.4°and 272.6°,respectively,while one set was established previously atφ=65.50.Each set of SXR arrays consists of three arrays viewing the plasma poloidally,and hence can be used separately to obtain SXR images via the tomographic method.The sawtooth precursor oscillations are measured by T-SXRI,and the corresponding images of perturbative SXR signals are successfully reconstructed at these three toroidal positions,hence providing measurement of the 3D structure of precursor oscillations.The observed 3D structure is consistent with the helical structure of the m/n=1/1 mode.The experimental observation confirms that the T-SXRI system is able to observe 3D structures in the J-TEXT plasma.

Intraoperative application of three-dimensional printed guides in total hip arthroplasty: A systematic review

BACKGROUND Acetabular component positioning in total hip arthroplasty(THA)is of key importance to ensure satisfactory post-operative outcomes and to minimize the risk of complications.The majority of acetabular components are aligned freehand,without the use of navigation methods.Patient specific instruments(PSI)and three-dimensional(3D)printing of THA placement guides are increasingly used in primary THA to ensure optimal positioning.AIM To summarize the literature on 3D printing in THA and how they improve acetabular component alignment.METHODS PubMed was used to identify and access scientific studies reporting on different 3D printing methods used in THA.Eight studies with 236 hips in 228 patients were included.The studies could be divided into two main categories;3D printed models and 3D printed guides.RESULTS 3D printing in THA helped improve preoperative cup size planning and post-operative Harris hip scores between intervention and control groups(P=0.019,P=0.009).Otherwise,outcome measures were heterogeneous and thus difficult to compare.The overarching consensus between the studies is that the use of 3D guidance tools can assist in improving THA cup positioning and reduce the need for revision THA and the associated costs.CONCLUSION The implementation of 3D printing and PSI for primary THA can significantly improve the positioning accuracy of the acetabular cup component and reduce the number of complications caused by malpositioning.

Investigation of the Acceptability of Cultured Meat in University Students

Background: Over the past 20 years, cultured meat has drawn a lot of public attention as a potential solution to issues with animal husbandry, including inadequate use of natural sources, improper animal welfare practices, and possible risks to public health and safety. The novel method of producing meat through culture reduces the need for animals to produce muscle fiber, thereby obviating the necessity for animal slaughter. Apart from its ethical advantages, cultured meat presents a possible way to fulfill the expanding need for food among growing populations. The purpose of this research was to find out whether Turkish students would be willing to pay for and accept cultured meat. Technique: Method: 371 university students who willingly consented to fill out a questionnaire and provide demographic data make up the research sample. Questions from previous studies on the acceptability of cultured meat were compiled to create the survey. The research’s data collection took place in March and April of 2022. The research was completed in June 2022 after the data had been processed and analyzed. Results: The results showed that the majority of participants were female and had omnivorous eating habits. Based on the results of the Bonferroni correction test, students with a higher intention to purchase and consume cultured meat were those who received economics and business education. Students with two years of university education had a higher overall survey score than those with four years of education (p < 0.05). Furthermore, it is discovered that there is a negative correlation between the participants’ ages and their Factor 2 (using cultured meat as an alternative to industrial meat) and Factor 3 (consuming and purchasing it) section points (r = -109, p = 0.036) (r = -0.121, p = 0.019). In conclusion, university students generally have a negative outlook on health-related issues, such as eating cultured meat as an alternative.

Computer-assisted three-dimensional individualized extreme liver resection for hepatoblastoma in proximity to the major liver vasculature

BACKGROUND The management of hepatoblastoma(HB)becomes challenging when the tumor remains in close proximity to the major liver vasculature(PMV)even after a full course of neoadjuvant chemotherapy(NAC).In such cases,extreme liver resection can be considered a potential option.AIM To explore whether computer-assisted three-dimensional individualized extreme liver resection is safe and feasible for children with HB who still have PMV after a full course of NAC.METHODS We retrospectively collected data from children with HB who underwent surgical resection at our center from June 2013 to June 2023.We then analyzed the detailed clinical and three-dimensional characteristics of children with HB who still had PMV after a full course of NAC.RESULTS Sixty-seven children diagnosed with HB underwent surgical resection.The age at diagnosis was 21.4±18.8 months,and 40 boys and 27 girls were included.Fifty-nine(88.1%)patients had a single tumor,39(58.2%)of which was located in the right lobe of the liver.A total of 47 patients(70.1%)had PRE-TEXT III or IV.Thirty-nine patients(58.2%)underwent delayed resection.After a full course of NAC,16 patients still had close PMV(within 1 cm in two patients,touching in 11 patients,compressing in four patients,and showing tumor thrombus in three patients).There were 6 patients of tumors in the middle lobe of the liver,and four of those patients exhibited liver anatomy variations.These 16 children underwent extreme liver resection after comprehensive preoperative evaluation.Intraoperative procedures were performed according to the preoperative plan,and the operations were successfully performed.Currently,the 3-year event-free survival of 67 children with HB is 88%.Among the 16 children who underwent extreme liver resection,three experienced recurrence,and one died due to multiple metastases.CONCLUSION Extreme liver resection for HB that is still in close PMV after a full course of NAC is both safe and feasible.This approach not only reduces the necessity for liver transplantation but also results in a favorable prognosis.Individualized three-dimensional surgical planning is beneficial for accurate and complete resection of HB,particularly for assessing vascular involvement,remnant liver volume and anatomical variations.

Three-dimensional visualization technology for guiding one-step percutaneous transhepatic cholangioscopic lithotripsy for the treatment of complex hepatolithiasis

BACKGROUND Biliary stone disease is a highly prevalent condition and a leading cause of hospitalization worldwide.Hepatolithiasis with associated strictures has high residual and recurrence rates after traditional multisession percutaneous transhepatic cholangioscopic lithotripsy(PTCSL).AIM To study one-step PTCSL using the percutaneous transhepatic one-step biliary fistulation(PTOBF)technique guided by three-dimensional(3D)visualization.METHODS This was a retrospective,single-center study analyzing,140 patients who,between October 2016 and October 2023,underwent one-step PTCSL for hepatolithiasis.The patients were divided into two groups:The 3D-PTOBF group and the PTOBF group.Stone clearance on choledochoscopy,complications,and long-term clearance and recurrence rates were assessed.RESULTS Age,total bilirubin,direct bilirubin,Child-Pugh class,and stone location were similar between the 2 groups,but there was a significant difference in bile duct strictures,with biliary strictures more common in the 3D-PTOBF group(P=0.001).The median follow-up time was 55.0(55.0,512.0)days.The immediate stone clearance ratio(88.6%vs 27.1%,P=0.000)and stricture resolution ratio(97.1%vs 78.6%,P=0.001)in the 3D-PTOBF group were significantly greater than those in the PTOBF group.Postoperative complication(8.6%vs 41.4%,P=0.000)and stone recurrence rates(7.1%vs 38.6%,P=0.000)were significantly lower in the 3D-PTOBF group.CONCLUSION Three-dimensional visualization helps make one-step PTCSL a safe,effective,and promising treatment for patients with complicated primary hepatolithiasis.The perioperative and long-term outcomes are satisfactory for patients with complicated primary hepatolithiasis.This minimally invasive method has the potential to be used as a substitute for hepatobiliary surgery.