Biology subjects

Yousef, H.

Publications and source records attributed to Yousef, H..

5 recordsLinked to original sources

bioRxiv2025

Blockade of VCAM1 or VLA4 preserves cerebrovasculature and prevents cognitive decline late after stroke

Infarct-induced neurodegeneration occurs chronically after stroke, doubling the risk of dementia. Endothelial vascular cell adhesion molecule 1 (VCAM1) facilitates blood-brain barrier opening and immune cell diapedesis by binding very late antigen 4 (VLA4) on immune cells. We hypothesized that vascular dysfunction persists after stroke and contributes to chronic neuroinflammation and cognitive decline via signaling through the VLA4/VCAM1 axis. We used adult (3-5 month old) and middle-aged (10 month old) C57BL/6J male & female mice and a permanent middle cerebral artery occlusion stroke model. Sham surgery consisted of an identical procedure without occlusion of the artery. We quantified vascular integrity using blood vessel length, pericyte coverage of vasculature, tight junctions, and extravascular fibrinogen leakage by immunostaining. Cognitive testing was performed using both Barnes maze and novel object recognition prior to stroke, and 1 and 6 weeks after stroke, and replicated in both male and female mice. We utilized anti-VCAM1, anti-VLA4 or isotype control antibodies to block VCAM1 or VLA4 function, and then to confirm mechanisms we utilized single cell RNA sequencing on immune and endothelial cells, aptamer-based plasma proteomics, and additional immunostaining for vascular integrity. Mouse brains exhibited signs of persistent vascular dysfunction and loss of blood-brain barrier integrity at 8 weeks after stroke, compared to sham animals. We observed reduced ZO-1 tight junction and pericyte coverage of vasculature, and increased extravascular fibrinogen. Mice with stroke also developed a cognitive deficit in both Barnes maze and novel object by 6 weeks. Treatment with anti-VCAM1 or anti-VLA4 resulted in mice with stroke performing as well as sham mice treated with isotype control antibody on both the Barnes maze and novel object recognition tasks. Anti-VCAM1 and anti-VLA4 both increased expression of blood-brain barrier maintenance genes in brain endothelial cells, while only minimally altering immune cell gene expression. Immune cell infiltration was reduced by anti-VCAM1 but not anti-VLA4 in tissue sections. In contrast to this, both antibodies increased blood vessel length and pericyte vascular coverage. Finally, extravascular fibrinogen was reduced by both antibody treatments in multiple brain regions. Together, our findings establish the VLA4/VCAM1 axis as a promising target to preserve vascular integrity and prevent cognitive decline late after stroke. Our data is consistent with a model where blocking either VCAM1 or VLA4 chronically after stroke promotes new blood vessel growth and maturation and restores the blood-brain barrier to prevent infarct-induced neurodegeneration.

neuroscience↗

bioRxiv2022

Observation of a single protein by ultrafast X-ray diffraction

The idea of using ultrashort X-ray pulses to obtain images of single proteins frozen in time has fascinated and inspired many. It was one of the arguments for building X-ray free-electron lasers. According to theory1, the extremely intense pulses provide sufficient signal to dispense with using crystals as an amplifier, and the ultrashort pulse duration permits capturing the diffraction data before the sample inevitably explodes2. This was first demonstrated on biological samples a decade ago on the giant mimivirus3. Since then a large collaboration4 has been pushing the limit of the smallest sample that can be imaged5,6. The ability to capture snapshots on the timescale of atomic vibrations, while keeping the sample at room temperature, may allow probing the entire conformational phase space of macromolecules. Here we show the first observation of an X-ray diffraction pattern from a single protein, that of Escherichia coli GroEL which at 14 nm in diameter7 is the smallest biological sample ever imaged by X-rays, and demonstrate that the concept of diffraction before destruction extends to single proteins. From the pattern, it is possible to determine the approximate orientation of the protein. Our experiment demonstrates the feasibility of ultrafast imaging of single proteins, opening the way to single-molecule time-resolved studies on the femtosecond timescale.

biophysics↗

bioRxiv2019

Undulating changes in human plasma proteome across lifespan are linked to disease

Aging is the predominant risk factor for numerous chronic diseases that limit healthspan. Mechanisms of aging are thus increasingly recognized as therapeutic targets. Blood from young mice reverses aspects of aging and disease across multiple tissues, pointing to the intriguing possibility that age-related molecular changes in blood can provide novel insight into disease biology. We measured 2,925 plasma proteins from 4,331 young adults to nonagenarians and developed a novel bioinformatics approach which uncovered profound non-linear alterations in the human plasma proteome with age. Waves of changes in the proteome in the fourth, seventh, and eighth decades of life reflected distinct biological pathways, and revealed differential associations with the genome and proteome of age-related diseases and phenotypic traits. This new approach to the study of aging led to the identification of unexpected signatures and pathways of aging and disease and offers potential pathways for aging interventions.

systems biology↗

bioRxiv2019

Brain endothelial cells are exquisite sensors of age-related circulatory cues

Brain endothelial cells (BECs) are key elements of the blood-brain barrier (BBB), protecting the brain from pathogens and restricting access to circulatory factors. Recent studies have demonstrated that the circulatory environment can modulate brain aging, yet, the underlying processes remain largely unknown. Given the BBBs intermediary position, we hypothesized that BECs sense, adapt to, and relay signals between the aging blood and brain. We sequenced single endothelial cells from the hippocampus--a brain region key to learning, memory, and neurogenesis-- of healthy young and aged mice as well as post-exposure to inflammatory and age-related circulatory factors. We discovered that aged capillary BECs, compared with arterial and venous cells, exhibit the greatest transcriptional changes, upregulating innate immunity, antigen presentation, TGF-{beta} signaling and oxidative stress response pathways. Remarkably, short-term infusions of aged plasma into young mice recapitulated key aspects of this aging transcriptome, while infusions of young plasma into aged mice reversed select aging signatures, essentially rejuvenating the BBB endothelium transcriptome. We identify candidate pathways mediating blood-borne brain rejuvenation by comparing age-upregulated genes with those modulated by plasma exposure. Together, these findings suggest that the transcriptional age of BECs is exquisitely sensitive to age-related circulatory cues and pinpoint the BBB itself as a promising therapeutic target to treat brain disease.\n\nHighlightsO_LISingle-cell RNA sequencing of brain endothelial cells (BECs) reveals transcriptional segmentation into distinct arterial, capillary, and venous identities with age and experimental interventions\nC_LIO_LIChanges with age are heterogenous across vessel segments, with aged capillaries enriched in signatures of innate immunity, TGF-{beta} and VEGF signaling, hypoxia and oxidative stress\nC_LIO_LIBECs sense and respond transcriptionally to diverse circulatory cues: inflammatory, proaging, or rejuvenating\nC_LIO_LIAged plasma exposure recapitulates--and young plasma reverses--key transcriptomic signatures of normal BEC aging\nC_LIO_LIBEC response to aged and young plasma reveals cell non-autonomous mechanisms of blood-brain-barrier aging\nC_LI

neuroscience↗

bioRxiv2018

Aged blood inhibits hippocampal neurogenesis and activates microglia through VCAM1 at the blood-brain barrier

An aged circulatory environment can promote brain dysfunction and we hypothesized that the blood-brain barrier (BBB) mediates at least some of these effects. We observe brain endothelial cells (BECs) in the aged mouse hippocampus express an inflammatory transcriptional profile with focal upregulation of Vascular Cell Adhesion Molecule 1 (VCAM1), a protein that facilitates vascular-immune cell interactions. Concomitantly, the shed, soluble form of VCAM1 is prominently increased in the aged circulation of humans and mice, and aged plasma is sufficient to increase VCAM1 expression in cultured BECs and young mouse hippocampi. Systemic anti-VCAM1 antibody or genetic ablation of VCAM1 in BECs counteracts the detrimental effects of aged plasma on young brains and reverses aging aspects in old mouse brains. Thus, VCAM1 is a negative regulator of adult neurogenesis and inducer of microglial reactivity, establishing VCAM1 and the luminal side of the BBB as possible targets to treat age-related neurodegeneration.

neuroscience↗