Protocol to encapsulate cerebral organoids with alginate hydrogel shell to induce volumetric compression.

In vitro organoids, including cerebral organoids, are usually developed without mechanical compression, which may contribute to a delay in maturation. Here, we present a protocol for encapsulating cerebral organoids with a thin shell of low-concentration alginate hydrogel. We describe steps for organoid generation, microfluidic chip culture, Matrigel coating, expansion culture, and alginate encapsulation. We then detail procedures for maturation culture and organoid characterization. The moderate compressive stimulation that the shell provides promotes cell proliferation and neuronal maturation. For complete details on the use and execution of this protocol, please refer to Tang et al.1.

Electrostatic potential difference between tumor and paratumor regulates cancer stem cell behavior and prognose tumor spread.

Tumor spread is responsible for most deaths related to cancer. Increasing the accuracy of cancer prognosis is critical to reducing the high mortality rates in cancer patients. Here, we report that the electrostatic potential difference (EPD) between tumor and its paratumor tissue is a prognostic marker for tumor spread. This finding is concluded from the patient-specific EPD values and clinical observation. The electrostatic potential values were measured on tissue cryosections from 51 patients using Kelvin probe force microscopy (KPFM). A total of ~44% (15/34) patients of Vtumor-paratumor > 0 were featured with tumor spread, whereas only ~18% (2/11) patients of Vtumor-paratumor < 0 had tumor spread. Next, we found the increased enrichment of cancer stem cells in paratumors with lower electrostatic potentials using immunofluorescence imaging, which suggested the attribution of tumor spread to the galvanotaxis of cancer stem cells (CSCs) toward lower potential. The findings were finally validated in breast and lung spheroid models composed of differentiated cancer cells and cancer stem cells at the ratio of 1:1 and embedded in Matrigel dopped with negative-, neutral- and positive-charged polymers and CSCs prefer to spread out of spheroids to lower electrostatic potential sites. This work may inspire the development of diagnostic and prognostic strategies targeting at tissue EPDs and CSCs for tumor therapy.

Vascularized Tumor Spheroid-on-a-Chip Model Verifies Synergistic Vasoprotective and Chemotherapeutic Effects.

Prolyl hydroxylases (PHD) inhibitors have been observed to improve drug distribution in mice tumors via blood vessel normalization, increasing the effectiveness of chemotherapy. These effects are yet to be demonstrated in human cell models. Tumor spheroids are three-dimensional cell clusters that have demonstrated great potential in drug evaluation for personalized medicine. Here, we used a perfusable vascularized tumor spheroid-on-a-chip to simulate the tumor microenvironment in vivo and demonstrated that the PHD inhibitor dimethylallyl glycine prevents the degradation of normal blood vessels while enhancing the efficacy of the anticancer drugs paclitaxel and cisplatin in human esophageal carcinoma (Eca-109) spheroids. Our results point to the potential of this model to evaluate anticancer drugs under more physiologically relevant conditions.

Multiplex gene quantification as digital markers for extremely rapid evaluation of chemo-drug sensitivity.

Current administrations for precision drug uses are limited in evaluation speed. Here, we propose the use of multiplex gene-based digital markers for the extremely rapid personalized prediction of individual sensitivity to cancer drugs. We first screen the transcriptional profiles by applying two to three gene filters and scoring genes by their impact on drug sensitivity and finalize the gene lists by K-nearest neighbors cross-validation. The digital markers are cancer type dependent, are composed of tens to hundreds of gene expressions, and are rapidly quantified by reverse transcription quantitative real-time PCR (qRT-PCR) within 1-3 h after tumor sampling. The area under the receiver operating characteristic curve reached 0.88 when testing the performance of digital markers on organoids derived from colorectal cancer patient tumors. The algorithm and corresponding graphic user interface were developed to demonstrate the promise of digital markers for extremely rapid drug recommendation.

Injectable Mesenchymal Stem Cell-Laden Matrigel Microspheres for Endometrium Repair and Regeneration.

Endometrial injury and intrauterine adhesions are increasingly reported in recent years; however, treatment options remain limited. Intravenous injection of mesenchymal stem cells (MSCs) for endometrium regeneration has limited effectiveness as the retention rate of transplanted cells is low. Hydrogel-based tissue-engineered solutions, such as MSC-seeded bioscaffolds, are reported to increase retention rates; however, a less invasive alternative is still desirable. 560-µm homogeneous Matrigel microspheres are fabricated, loading them with about 1500 MSCs and injecting them into the injured endometria of rats' uteri. This minimally invasive procedure is proved to significantly increase endometrium thickness by over onefold after 21 d (p < 0.0001) and fertility rates from 25% to 75% in impaired and repaired uteri (p < 0.001), respectively. This study provides a minimally invasive alternative to endometrium repair with the promise to establish a broad-spectrum technique for MSC transplantation.

Integrating Engineering, Automation, and Intelligence to Catalyze the Biomedical Translation of Organoids.

Organoid technology has developed at an impressive speed during the past decade. Still, organoids are not widely used in practical applications as expected. It is believed that this translation can be greatly accelerated with the integration of engineering and artificial intelligence into current research practices. It is proposed that this approach is the missing link to realize key milestones in organoid technology, namely, high-throughput, homogeneous, and standardized production, automated manipulation, and intelligent monitoring, evaluation, and control via integrated on-chip instrumentation and artificial intelligence. It is suggested that organoids-on-a-chip are the ideal platform to achieve these feats. Once these techniques are established and adopted by the scientific community, the rapid translation of organoids may be seen from laboratories to the clinics and pharmaceutical industry.