Written By: Abel B. Cortinas
Written by: Ken Estrellas
Original Article: Hu et al. Biology Open 2019
The Gist of It:
Breast cancer is one of the most common cancers, with numerous cases reported worldwide every year. One of the earliest, localized stages of this disease, known as DCIS (ductal carcinoma in situ), represents about 1/5 of new breast cancer cases annually. Although the prognosis is positive, with 99% of patients surviving after 5 years and 95% surviving after 20 years, there is always a fear of the disease spreading beyond local tissue and beginning to metastasize. Understanding what drives disease progression in cancer is a topic of persistent interest. With that in mind, Dr. Hu and colleagues have demonstrated an interesting similarity between breast cancer cells and healthy macrophages, an important cell in the immune system – both types of cells respond positively to a protein known as CCL2. CCL2 is a chemokine, a type of protein that attracts certain types of cells to the tissues where it is being made. Normally, CCL2 binds to the receptor CCR2 to help attract macrophages in response to infections, inflammation, or other scenarios such as wound healing. In breast cancer, the presence of more CCL2 has been associated with disease progression. In this study, breast cancer cells that were modified to be even more responsive to CCL2 showed more invasive growth patterns, invasive potential, and expression of genes associated with tumor growth, along with reduced expression of a tumor suppressor gene. Notably, when breast cancer cells had the receptor CCR2 edited out, the cells were no longer responsive to CCL2, and their growth and invasion were inhibited when treated with CCL2. Additionally, expression of the gene ALDH1A1, which is known to be a pro-invasion factor for tumor cells, was associated with more CCR2 in the cells, more responsiveness to CCL2, and thus increased potential for invasion and metastasis. On the other hand, expression of the gene HTRA2, which is known to play a role in cell death, was associated with the opposite effect. Taken together, these findings paint a clearer picture of the mechanisms behind breast cancer metastasis and could someday help physicians diagnose and detect potential metastasis earlier in the disease cycle.
The Nitty Gritty
In this study, Hu et al. elucidated the roles of various components of the CCL2/CCR2 signaling axis in the proliferation, invasion, and potential metastasis of breast cancer cells. The breast cancer cell line SUM225, which has reduced expression of CCR2, was less prone to asymmetric cell growth patterns in the presence of recombinant CCL2 compared with the DCIS.com cell line, which has greater expression of CCR2. SUM225 cells modified to overexpress CCR2 subsequently showed increased responsiveness to CCL2, indicated by greater spheroid size, increased mean inversion index, increased expression of PCNA and TWIST1, and decreased expression of E-cadherin. Conversely, CRISPR-Cas9–mediated ablation of CCR2 in DCIS.com cells eliminated this effect, rendering the cells less responsive to CCL2 as indicated by decreased spheroid formation, lower levels of invasion, and decreased PCNA expression. Expression of the metabolic enzyme ALDH1A1 was increased in SUM225 cells treated with CCL2 and decreased in DCIS.com cells in which CCR2 expression was reduced by shRNA knockdown or CRISPR knockout treated with CCL2, suggesting that CCR2 positively regulates ALDH1A1 expression in both cell lines. This was supported by the fact that ALDH1A1 overexpression and knockdown affected the growth of CCR2-overexpressing SUM225 cells and CCR2-knockout DCIS.com cells, but not their invasion. The converse effect was observed with expression of the pro-apoptotic mitochondrial serine protease HTRA2, suggesting that CCR2 negatively regulates HTRA2. Taken together, these results reinforce previous literature associating the CCL2/CCR2 signaling axis with breast cancer metastasis and suggest that ALDH1A1 may play a specific role in mediating this process.
Original Research Article: Hu Q., et al. “Role of ALDH1A1 and HTRA2 expression in CCL2/CCR2-mediated breast cancer cell growth and invasion.” Biol Open 8.7 (2019): pii: bio040873. doi: 10.1242/bio.040873.
Written By: Kaitlyn Sadtler
Original Article: Sharma & Subramaniam. Journal of Cell Biology 2019
The Gist of It:
Huntington’s disease is a heritable degenerative brain disorder that is caused by a mutant protein – called Huntingtin (HTT) – which has something called a “triplet repeat expansion.” The diseased, mutant version has a CAG triplet repeat that is expanded, which means that the DNA sequence “CAG” is mistakenly replicated many more times than it should be (40–50+ of these CAG repeats); this causes clumping of the proteins and the disease. The effects of this disease are both mental and physical, with symptoms ranging from involuntary movements and impaired balance to depression and anxiety. There is no cure, and patients will die from their illness within 15–20 years of the initial onset of symptoms, which usually occurs from ages 30–50. Recently, researchers have discovered that the diseased protein can spread to neighboring brain cells through tunnel-like passages that form between a “donor cell” and an “acceptor cell”. This process depends upon the action of another protein called Rhes. In a set of experiments, the researchers were able to show that the protein Rhes creates small tunnels between cells that can transport mutant HTT, but not normal HTT. Also, the protein Rhes can tag things with a small protein called “SUMO” (think of this like putting a post-it note on a page of a book). If Rhes is genetically modified so it can no longer put its SUMO post-it note on mutant HTT, then the transport of mutant HTT from one cell to another is decreased. Figuring out how these mutant proteins spread in the human body can help identify new targets for treating diseases such as Huntington’s by stopping further progression or even ameliorating existing symptoms.
The Nitty Gritty:
Previous research has identified the presence of tunneling nanotubes (TNTs) that provide cytoplasmic connections between neighboring cells. In this study, Sharma and Subramaniam describe TNT-like structures that are capable of transporting mutant Huntingtin protein (mHTT) but not wild-type HTT from a donor cell to an acceptor cell. This transfer is dependent upon the GTPase/SUMO E3-like protein Rhes, which is found mainly in brain tissue. When transfected into striatal cells (STHdhQ7/Q7), GFP-linked Rhes induced the formation of filipodia-like protrusions, which were not present in cells transfected with GFP-RhoA or GFP alone. These protrusions connected with other cells transiently in a “kiss-and-run” fashion wherein membrane fusion was detected via scanning and transmission electron microscopy. This behavior was prevented with the addition of the actin polymerization inhibitor cytochalasin D. The transition of Rhes between cells with TNT-like connections was shown by tagging Rhes with GFP and acceptor cell membranes with mCherry and measuring double-positive cells via flow cytometry. TNT-like Rhes tunnels are capable of transiting mHTT between neighboring cells but not wild type HTT or other proteins such as mTOR or wild type Tau. If the SUMOylation activity of Rhes was inhibited (Rhes C263S), mHTT SUMOylation was inhibited, or SUMO was knocked out using CRISPR/Cas9, transport of mHTT was significantly inhibited. These findings describe a form of transfer of mHTT from neighboring cells through direct connection of the cytoplasms via Rhes-dependent TNT-like connections.
Original Research Article: Sharma, M. & Subramaniam, S. “Rhes travels from cell to cell and transports Huntington disease protein via TNT-like protrusion.” J Cell Biol 218.6 (2019): 1972.
Written By: Rebecca Tweedell
Original Article: Rodgers et al. Antimicrobial Agents and Chemotherapy 2019
The Gist of It:
Bacterial infections are a major threat to newborn babies. In developed countries, improved sanitation, monitoring, and treatment have greatly reduced the risk of infection, but in low- and middle-income countries, bacterial infections can still be a death sentence for a baby. One of the critical antibiotics used to combat these bacterial infections is gentamicin because it is cheap and effective. Unfortunately, it is normally given as a shot into the muscle (like the flu vaccine is), and this requires trained medical workers with access to safe needle disposal, which is not always available in the areas where the drug is needed the most. To avoid the drawbacks of gentamicin, researchers have been working on finding new ways to get the drug into people’s bodies. Researchers at Queens University Belfast in the United Kingdom recently invented a microarray patch that can deliver gentamicin without the need for shots. The microarray patch looks similar to the nicotine patches that you may have seen, and it uses short, pain-free, microscopic needles that dissolve to release the gentamicin. This allows the microarray patch to be used by healthcare workers without the need for special training and needle disposal. The researchers put these patches to the test in a mouse model of Klebsiella pneumoniae infection; K. pneumoniae is a bacterium that can cause pneumonia, bloodstream infections, and intra-abdominal infections in humans. Mice were infected with K. pneumoniae and then treated with a traditional shot of gentamicin or the microarray patch. The researchers found that both treatments led to significant decreases in the number of bacteria in the lungs and lymphoid tissue in the mice. This means that microarray patches could potentially be used in place of gentamicin shots, reducing the need for needles, needle disposal, and trained staff. Similar methods could also be used to improve the delivery of other treatments, vastly improving access to medical care for those in remote and poor areas and saving the lives of thousands of infants.
The Nitty Gritty:
Microarray patches containing gentamicin were fabricated in two parts, the baseplate and the needles. Baseplates were made from 15% polyvinylpyrrolidone (PVP) cast into a mold to create a thin rectangular cuboid shape. The microscopic needles were made by adding a solution containing 3.4% sodium hyaluronate, 1% PVP, and 10% gentamicin sulfate to a sawtooth-shaped mold and applying 3–4 bar pressure for 15 minutes to push the mix into the grooves of the mold. The baseplate was then applied to the base of the needles, and the entire assembly was dried for 48 hours before the full microarray plate was removed from the mold. Each resulting needle was approximately 500 μm long and 300 μm wide at the base. To test the efficacy of the microarray plate in vivo, C57BL6 mice were inoculated with K. pneumoniae intranasally. Eight hours after infection, mice were treated with an intramuscular injection of gentamicin (7.5 mg/kg body weight), the gentamicin microarray plate (placed on the ear and secured with micropore tape, then replaced with a fresh microarray plate at 24 hours post-infection), or no treatment. Bacterial levels in the nasal-associated lymphoid tissue (NALT), lungs, and spleens were quantified 48 hours post-infection. Mice treated with the microarray plate had statistically significant reductions in bacterial levels in the NALT and lungs compared with bacterial levels in untreated mice. Treatment with the intramuscular injection of gentamicin produced similar results. These results showed in vivo evidence that the microarray plate could be used as an alternative delivery method for life-saving gentamicin treatment.
Original Research Article: Rodgers, A.M., et al. “Control of Klebsiella pneumoniae infection in mice by using dissolving microarray patches containing gentamicin.” Antimicrob Agents Chemother 63 (2019): e02612-18.
Written By: Kaitlyn Sadtler