Clinical efficacy and safety of robotic retroperitoneal lymph node dissection for testicular cancer: a systematic review and meta-analysis.

Background: Retroperitoneal lymph node dissection (RPLND) is an effective treatment for testicular tumors. In recent years, with the development of robotics, many urological procedures performed via standard laparoscopy have been replaced by robots. Our objective was to compare the safety and efficacy of robotic retroperitoneal lymph node dissection (R-RPLND) versus Non-robotic retroperitoneal lymph node dissection (NR-RPLND) in testicular cancer. Methods: Pubmed, Embase, Scopus, Cochrane Library, and Web of Science databases were searched for literature on robotic surgery for testicular germ cell tumors up to April 2023. The statistical and sensitivity analyses were performed using Review Manager 5.3. Meta-analysis was performed to calculate mean difference (MD), odds ratio(OR), and 95% confidence interval (CI) effect indicators. Results: Eight studies with 3875 patients were finally included in this study, 453 with R-RPLND and 3422 with open retroperitoneal lymph node dissection (O-RPLND)/laparoscopic retroperitoneal lymph node dissection (L-RPLND). The results showed that R-RPLND had lower rates of intraoperative blood loss (MD = -436.39; 95% CI -707.60 to -165.19; P = 0.002), transfusion (OR = 0.06; 95% CI 0.01 to 0.26; P = 0.0001), total postoperative complication rates (OR = 0.39; 95% CI 0.21 to 0.70; P = 0.002), and length of stay (MD=-3.74; 95% CI -4.69 to -2.78; P<0.00001). In addition, there were no statistical differences between the two groups regarding perioperative and oncological outcomes regarding total operative time, the incidence of postoperative complications grade≥III, abnormal ejaculation rate, lymph node yield, and postoperative recurrence rate. Conclusions: The R-RPLND and O-RPLND/L-RPLND provide safe and effective retroperitoneal lymph node dissection for testicular cancer. Patients with R-RPLND have less intraoperative bleeding, shorter hospitalization period, fewer postoperative complications, and faster recovery. It should be considered a viable alternative to O-RPLND/L-RPLND. Systematic Review Registration: https://www.crd.york.ac.uk/PROSPERO, identifier CRD42023411696.

Molecular and functional characterization of MST2 in grass carp during bacterial infection.

The regulation of host redox homeostasis is critically important in the immune response to pathogens. The "mammalian sterile 20-like" kinase 2 (MST2) has been shown to play a role in apoptosis, cell proliferation, and cancer; however, few studies have examined its ability to modulate redox homeostasis during innate immunity, especially in teleost fish. In this study, we cloned the MST2 gene of Ctenopharyngodon idella (CiMST2) and analyzed its tissue distribution. CiMST2 was present in most of the studied tissues, and it was most highly expressed in brain tissue. Expression patterns analysis revealed that MST2 mRNA and protein were significantly up-regulated under bacterial infection, suggesting that it is involved in the immune response. Bacterial stimulation significantly increased the level of antioxidases. To explore the interplay between CiMST2 and antioxidant regulation, we examined the effects of CiMST2 overexpression and conducted RNA interference assays in vitro. CiMST2 overexpression significantly increased the expression levels of nuclear factor E2-related factor 2 (Nrf2) and other antioxidases and vice versa, revealing that CiMST2 regulated host redox homeostasis via Nrf2-antioxidant responsive element (ARE) signaling. Overall, our findings provide a new perspective on the role of MST2 in innate immunity in teleosts as well as insights that will aid the prevention and control of disease in the grass carp farming industry.

Introgressing the Aegilops tauschii genome into wheat as a basis for cereal improvement.

Increasing crop production is necessary to feed the world's expanding population, and crop breeders often utilize genetic variations to improve crop yield and quality. However, the narrow diversity of the wheat D genome seriously restricts its selective breeding. A practical solution is to exploit the genomic variations of Aegilops tauschii via introgression. Here, we established a rapid introgression platform for transferring the overall genetic variations of A. tauschii to elite wheats, thereby enriching the wheat germplasm pool. To accelerate the process, we assembled four new reference genomes, resequenced 278 accessions of A. tauschii and constructed the variation landscape of this wheat progenitor species. Genome comparisons highlighted diverse functional genes or novel haplotypes with potential applications in wheat improvement. We constructed the core germplasm of A. tauschii, including 85 accessions covering more than 99% of the species' overall genetic variations. This was crossed with elite wheat cultivars to generate an A. tauschii-wheat synthetic octoploid wheat (A-WSOW) pool. Laboratory and field analysis with two examples of the introgression lines confirmed its great potential for wheat breeding. Our high-quality reference genomes, genomic variation landscape of A. tauschii and the A-WSOW pool provide valuable resources to facilitate gene discovery and breeding in wheat.

Microfluidic filter device with nylon mesh membranes efficiently dissociates cell aggregates and digested tissue into single cells.

Tissues are increasingly being analyzed at the single cell level in order to characterize cellular diversity and identify rare cell types. Single cell analysis efforts are greatly limited, however, by the need to first break down tissues into single cell suspensions. Current dissociation methods are inefficient, leaving a significant portion of the tissue as aggregates that are filtered away or left to confound results. Here, we present a simple and inexpensive microfluidic device that simultaneously filters large tissue fragments and dissociates smaller aggregates into single cells, thereby improving single cell yield and purity. The device incorporates two nylon mesh membranes with well-defined, micron-sized pores that operate on aggregates of different size scales. We also designed the device so that the first filtration could be performed under tangential flow to minimize clogging. Using cancer cell lines, we demonstrated that aggregates were effectively dissociated using high flow rates and pore sizes that were smaller than a single cell. However, pore sizes that were less than half the cell size caused significant damage. We then improved results by passing the sample through two filter devices in series, with single cell yield and purity predominantly determined by the pore size of the second membrane. Next, we optimized performance using minced and digested murine kidney tissue samples, and determined that the combination of 50 and 15 µm membranes was optimal. Finally, we integrated these two membranes into a single filter device and performed validation experiments using minced and digested murine kidney, liver, and mammary tumor tissue samples. The dual membrane microfluidic filter device increased single cell numbers by at least 3-fold for each tissue type, and in some cases by more than 10-fold. These results were obtained in minutes without affecting cell viability, and additional filtering would not be required prior to downstream applications. In future work, we will create complete tissue analysis platforms by integrating the dual membrane microfluidic filter device with additional upstream tissue processing technologies, as well as downstream operations such as cell sorting and detection.

Microfluidic channel optimization to improve hydrodynamic dissociation of cell aggregates and tissue.

Maximizing the speed and efficiency at which single cells can be liberated from tissues would dramatically advance cell-based diagnostics and therapies. Conventional methods involve numerous manual processing steps and long enzymatic digestion times, yet are still inefficient. In previous work, we developed a microfluidic device with a network of branching channels to improve the dissociation of cell aggregates into single cells. However, this device was not tested on tissue specimens, and further development was limited by high cost and low feature resolution. In this work, we utilized a single layer, laser micro-machined polyimide film as a rapid prototyping tool to optimize the design of our microfluidic channels to maximize dissociation efficiency. This resulted in a new design with smaller dimensions and a shark fin geometry, which increased recovery of single cells from cancer cell aggregates. We then tested device performance on mouse kidney tissue, and found that optimal results were obtained using two microfluidic devices in series, the larger original design followed by the new shark fin design as a final polishing step. We envision our microfluidic dissociation devices being used in research and clinical settings to generate single cells from various tissue specimens for diagnostic and therapeutic applications.

Phenotypic Analysis of Stromal Vascular Fraction after Mechanical Shear Reveals Stress-Induced Progenitor Populations.

BACKGROUND: Optimization of fat grafting continues to gain increasing attention in the field of regenerative medicine. "Nanofat grafting" implements mechanical emulsification and injection of standard lipoaspirate for the correction of superficial rhytides and skin discoloration; however, little is known about the cellular constituents of the graft. Based on recent evidence that various stressors can induce progenitor activity, the authors hypothesized that the shear forces used in common fat grafting techniques may impact their regenerative capacities. METHODS: Lipoaspirates were obtained from 10 patients undergoing elective procedures. Half of each sample was subjected to nanofat processing; the other half was left unchallenged. The viscosity of each sample was measured for computational analysis. The stromal vascular fraction of each sample was isolated, quantified, and analyzed by means of flow cytometry with two multicolor fluorescence antibody panels. RESULTS: Standard lipoaspirate is ideally suited for mechanical stress induction. The mechanical emulsification involved in nanofat processing did not affect cell number; however, viability was greatly reduced when compared with the stromal vascular fraction of standard lipoaspirate. Interestingly, nanofat processing resulted in stress-induced stromal vascular fraction with a higher proportion of endothelial progenitor cells, mesenchymal stem cells, and multilineage differentiating stress-enduring cells. Single-parameter analysis also revealed significant increases in CD34, CD13, CD73, and CD146 of the stress-induced stromal vascular fraction, markers associated with mesenchymal stem cell activity. CONCLUSIONS: Mechanical processing used in techniques such as nanofat grafting induces the up-regulation of progenitor phenotypes consistent with multipotency and pluripotency. These data provide a first step in characterizing the potential regenerative benefits realized through stress induction in fat grafting. CLINCAL QUESTION/LEVEL OF EVIDENCE: Therapeutic, V.

Synthesis, molecular modeling, and biological evaluation of novel RAD51 inhibitors.

RAD51 recombinase plays a critical role for cancer cell proliferation and survival. Targeting RAD51 is therefore an attractive strategy for treating difficult-to-treat cancers, e.g. triple negative breast cancers which are often resistant to existing therapeutics. To this end, we have designed, synthesized and evaluated a panel of new RAD51 inhibitors, denoted IBR compounds. Among these compounds, we have identified a novel small molecule RAD51 inhibitor, IBR120, which exhibited a 4.8-fold improved growth inhibition activity in triple negative human breast cancer cell line MBA-MD-468. IBR120 also inhibited the proliferation of a broad spectrum of other cancer cell types. Approximately 10-fold difference between the IC50 values in normal and cancer cells were observed. Moreover, IBR120 was capable of disrupting RAD51 multimerization, impairing homologous recombination repair, and inducing apoptotic cell death. Therefore, these novel RAD51 inhibitors may serve as potential candidates for the development of pharmaceutical strategies against difficult-to-treat cancers.

Microfluidic device for mechanical dissociation of cancer cell aggregates into single cells.

Tumors tissues house a diverse array of cell types, requiring powerful cell-based analysis methods to characterize cellular heterogeneity and identify rare cells. Tumor tissue is dissociated into single cells by treatment with proteolytic enzymes, followed by mechanical disruption using vortexing or pipetting. These procedures can be incomplete and require significant time, and the latter mechanical treatments are poorly defined and controlled. Here, we present a novel microfluidic device to improve mechanical dissociation of digested tissue and cell aggregates into single cells. The device design includes a network of branching channels that range in size from millimeters down to hundreds of microns. The channels also contain flow constrictions that generate well-defined regions of high shear force, which we refer to as "hydrodynamic micro-scalpels", to progressively disaggregate tissue fragments and clusters into single cells. We show using in vitro cancer cell models that the microfluidic device significantly enhances cell recovery in comparison to mechanical disruption by pipetting and vortexing after digestion with trypsin or incubation with EDTA. Notably, the device enabled superior results to be obtained after shorter proteolytic digestion times, resulting in fully viable cells in less than ten minutes. The device could also be operated under enzyme-free conditions that could better maintain expression of certain surface markers. The microfluidic format is advantageous because it enables application of well-defined mechanical forces and rapid processing times. Furthermore, it may be possible to directly integrate downstream processing and detection operations to create integrated cell-based analysis platforms. The enhanced capabilities enabled by our novel device may help promote applications of single cell detection and purification techniques to tumor tissue specimens, advancing the current understanding of cancer biology and enabling molecular diagnostics in clinical settings.

A novel small molecule RAD51 inactivator overcomes imatinib-resistance in chronic myeloid leukaemia.

RAD51 recombinase activity plays a critical role for cancer cell proliferation and survival, and often contributes to drug-resistance. Abnormally elevated RAD51 function and hyperactive homologous recombination (HR) rates have been found in a panel of cancers, including breast cancer and chronic myeloid leukaemia (CML). Directly targeting RAD51 and attenuating the deregulated RAD51 activity has therefore been proposed as an alternative and supplementary strategy for cancer treatment. Here we show that a newly identified small molecule, IBR2, disrupts RAD51 multimerization, accelerates proteasome-mediated RAD51 protein degradation, reduces ionizing radiation-induced RAD51 foci formation, impairs HR, inhibits cancer cell growth and induces apoptosis. In a murine imatinib-resistant CML model bearing the T315I Bcr-abl mutation, IBR2, but not imatinib, significantly prolonged animal survival. Moreover, IBR2 effectively inhibits the proliferation of CD34(+) progenitor cells from CML patients resistant to known BCR-ABL inhibitors. Therefore, small molecule inhibitors of RAD51 may suggest a novel class of broad-spectrum therapeutics for difficult-to-treat cancers.

Synthesis and biological evaluation of a series of novel inhibitor of Nek2/Hec1 analogues.

High expression in cancer 1 (Hec1) is an oncogene overly expressed in many human cancers. Small molecule inhibitor of Nek2/Hec1 (INH) targeting the Hec1 and its regulator, Nek2, in the mitotic pathway, was identified to inactivate Hec1/Nek2 function mediated by protein degradation that subsequently leads to chromosome mis-segregation and cell death. To further improve the efficacy of INH, a series of INH analogues were designed, synthesized, and evaluated. Among these 33 newly synthesized analogues, three of them, 6, 13, and 21, have 6-8 fold more potent cell killing activity than the previous lead compound INH1. Compounds 6 and 21 were chosen for analyzing the underlying action mechanism. They target directly the Hec1/Nek2 pathway and cause chromosome mis-alignment as well as cell death, a mechanism similar to that of INH1. This initial exploration of structural/functional relationship of INH may advance the progress for developing clinically applicable INH analogue.

Small molecule targeting the Hec1/Nek2 mitotic pathway suppresses tumor cell growth in culture and in animal.

Hec1 is a conserved mitotic regulator critical for spindle checkpoint control, kinetochore functionality, and cell survival. Overexpression of Hec1 has been detected in a variety of human cancers and is linked to poor prognosis of primary breast cancers. Through a chemical genetic screening, we have identified a small molecule, N-(4-[2,4-dimethyl-phenyl]-thiazol-2-yl)-benzamide (INH1), which specifically disrupts the Hec1/Nek2 interaction via direct Hec1 binding. Treating cells with INH1 triggered reduction of kinetochore-bound Hec1 as well as global Nek2 protein level, consequently leading to metaphase chromosome misalignment, spindle aberrancy, and eventual cell death. INH1 effectively inhibited the proliferation of multiple human breast cancer cell lines in culture (GI(50), 10-21 micromol/L). Furthermore, treatment with INH1 retarded tumor growth in a nude mouse model bearing xenografts derived from the human breast cancer line MDA-MB-468, with no apparent side effects. This study suggests that the Hec1/Nek2 pathway may serve as a novel mitotic target for cancer intervention by small compounds.