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Biology subjects

Dallas, M.

Publications and source records attributed to Dallas, M..

3 recordsLinked to original sources

Long-term maintenance of patient-specific characteristics in tumoroids from six cancer indications in a common base culture media system

Tumoroids, also known as cancer organoids, are patient-derived cancer cells grown as 3D, self-organized multicellular structures that maintain key characteristics (e.g., genotype, gene expression levels) of the tumor from which they originated. These models have emerged as valuable tools for studying tumor biology, cytotoxicity, and response of patient-derived cells to cancer therapies. However, the establishment and maintenance of tumoroids has historically been challenging, labor intensive, and highly variable from lab to lab, hindering their widespread use. Here, we characterize the establishment and/or expansion of colorectal, lung, head and neck, breast, pancreas, and endometrial tumoroids using the standardized, serum-free Gibco OncoPro Tumoroid Culture Medium. Newly derived tumoroid lines (n=20) were analyzed by targeted genomic profiling and RNA sequencing and were representative of tumor tissue samples. Tumoroid lines were stable for over 250 days in culture and freeze-thaw competent. Previously established tumoroid lines were also transitioned to OncoPro medium and exhibited, on average, similar growth rates and conserved donor-specific characteristics when compared to original media systems. Additionally, OncoPro medium was compatible with both embedded culture in extracellular matrix and growth in a suspension format for facile culture and scale up. An example application of these models for assessing the cytotoxicity of a natural killer cell line and primary natural killer cells over time and at various doses demonstrated the compatibility of these models with assays used in compound and cell therapy development. We anticipate that the standardization and versatility of this approach will have important benefits for basic cancer research, drug discovery, and personalized medicine and help make tumoroid models more accessible to the cancer research community.

cancer biology↗

Kv1.3 induced hyperpolarisation and Cav3.2-mediated calcium entry are required for efficientKaposi's sarcoma-associated herpesvirus lytic replication

Understanding the host factors critical for Kaposis sarcoma-associated herpesvirus (KSHV) lytic replication can identify new targets for therapeutic intervention. Using pharmacological and genetic silencing approaches, we reveal for the first time that KSHV requires a B cell expressed voltage-gated K+ channel, Kv1.3, to enhance lytic replication. We show that the KSHV replication and transcription activator (RTA) protein upregulates Kv1.3 expression, leading to enhanced K+ channel activity and hyperpolarisation of the B cell membrane. Enhanced Kv1.3 activity then promotes intracellular Ca2+ influx through Cav3.2, a T-type Ca2+ channel, leading to the Ca2+ driven nuclear localisation of NFAT and the subsequent NFAT1-responsive gene expression. Importantly, KSHV lytic replication and infectious virion production could be inhibited by both Kv1.3 and Cav3.2 blockers or through Kv1.3 silencing. These findings provide new mechanistic insight into the essential role of host ion channels during KSHV infection and highlight Kv1.3 and Cav3.2 as new druggable host factors that are key to the successful completion of KSHV lytic replication.

microbiology↗

The hypoxic ventilatory response is facilitated by the activation of Lkb1-AMPK signalling pathways downstream of the carotid bodies

We recently demonstrated that the role of the AMP-activated protein kinase (AMPK), a ubiquitously expressed enzyme that governs cell-autonomous metabolic homeostasis, has been extended to system-level control of breathing and thus oxygen and energy (ATP) supply to the body. Here we assess the contribution to the hypoxic ventilatory response (HVR) of two upstream kinases that govern the activities of AMPK. Lkb1, which activates AMPK in response to metabolic stress and CaMKK2 which mediates the alternative Ca2+-dependent mechanism of AMPK activation. HVRs remained unaffected in mice with global deletion of the CaMKK2 gene. By contrast, HVRs were markedly attenuated in mice with conditional deletion of LKB1 in catecholaminergic cells, including carotid body type I cells and brainstem respiratory networks. In these mice hypoxia evoked hypoventilation, apnoea and Cheyne-Stokes-like breathing, rather than hyperventilation. Attenuation of HVRs, albeit less severe, was also conferred in mice carrying [~]90% knockdown of Lkb1 expression. Carotid body afferent input responses were retained following either [~]90% knockdown of Lkb1 or AMPK deletion. In marked contrast, LKB1 deletion virtually abolished carotid body afferent discharge during normoxia, hypoxia and hypercapnia. We conclude that Lkb1 and AMPK, but not CaMKK2, facilitate HVRs at a site downstream of the carotid bodies.

physiology↗