Transit time ultrasound perivascular flow probe technology is superior to MR imaging on hepatic blood flow measurement in a porcine model

Background: The hepatic hemodynamics is an essential parameter in surgical planning as well as in various disease processes. The transit time ultrasound(TTUS) perivascular flow probe technology is widely used in clinical practice to evaluate the hepatic inflow, yet invasive. The phase-contrast-MRI(PC-MRI) is not invasive and potentially applicable in assessing the hepatic blood flow. In the present study, we compared the hepatic inflow rates using the PC-MRI and the TTUS probe, and evaluated their predictive value of post-hepatectomy adverse events. Methods: Eighteen large white pigs were anaesthetized for PC-MRI and approximately 75% hepatic resection was performed under a unified protocol. The blood flow was measured in the hepatic artery(Qha), the portal vein(Qpv), and the aorta above the celiac trunk(Qca) using PC-MRI, and was compared to the TTUS probe. The Bland-Altman method was conducted and a partial least squares regression(PLS) model was implemented. Results: The mean Qpv measured in PC-MRI was 0.55 ± 0.12 L/min, and in the TTUS probe was 0.74 ± 0.17 L/min. Qca was 1.40 ± 0.47 L/min in the PC-MRI and 2.00 ± 0.60 L/min in the TTUS probe. Qha was 0.17 ± 0.10 L/min in the PC-MRI, and 0.13 ± 0.06 L/min in the TTUS probe. The Bland-Altman method revealed that the estimated bias of Qca in the PC-MRI was 32%(95% CI:-49% to 15%); Qha 17%(95% CI:-15% to 51%); and Qpv 40%(95% CI:-62% to 18%). The TTUS probe had a higher weight in predicting adverse outcomes after 75% resection compared to the PC-MRI( β= 0.35 and 0.43 vs β = 0.22 and 0.07, for tissue changes and premature death, respectively). Conclusions: There is a tendency of the PC-MRI to underestimate the flow measured by the TTUS probes. The TTUS probe measures are more predictive of relevant post-hepatectomy outcomes.     

Deregulation of the expression of the fractalkine/fractalkine receptor complex in HIV-1-infected patients

Fractalkine is the only member of the CX3C chemokine family. Polymorphism of the fractalkine receptor gene may influence the prognosis of human immunodeficiency virus (HIV) infection, but the nature of the cells expressing fractalkine or its receptor in HIV-infected patients remains unknown. We show that, in contrast to HIV-uninfected individuals, a large number of cells expressed fractalkine in T-cell zones of lymph nodes from HIV-infected patients. CD83(+) mature and CD123(+) plasmacytoid dendritic cells as well as plasma cells are involved in this increased expression of fractalkine. Increased numbers of plasmacytoid dendritic cells and plasma cells were present in T-cell zones of HIV-infected patients. CD83(+) dendritic cells were present in similar number in HIV-infected patients and controls, but an increased fraction of these cells produced fractalkine in HIV-infected patients. Many plasma cells in the gut-associated lymphoid tissue from HIV-infected patients also produced fractalkine, whereas few cells produced fractalkine in the gut of controls. The fraction of CD45RO(+) and CD45RO(-) T helper (Th) cells expressing the fractalkine receptor CX3CR1 was higher in HIV-infected patients than in healthy individuals, and these cells were abnormally sensitive to fractalkine stimulation. This increased response correlated with HIV viremia, and it returned to normal levels in patients successfully treated with antiretroviral drugs. The increased expression of the fractalkine/fractalkine receptor complex associated with HIV infection may affect adhesion and migration of Th lymphocytes and their interaction with dendritic cells. Thus, it may influence the equilibrium between depletion and renewal of the Th lymphocyte compartment.     

Anastomotic salvage after rectal cancer resection using the Turnbull–Cutait delayed anastomosis

Background

Turnbull–Cutait abdominoperineal pull-through followed by delayed coloanal anastomosis (DCA) was first described in 1961. Studies have described its use for challenging colorectal conditions. We reviewed our experience with Turnbull–Cutait DCA as a salvage procedure for complex failure of colorectal anastomosis.

Methods

We performed a retrospective cohort study from October 2010 to September 2011, with analysis of postoperative morbidity and mortality.

Results

Seven DCAs were performed for anastomotic complications (3 chronic leaks, 2 rectovaginal fistulas, 1 colovesical fistula, 1 colonic ischemia) following surgery for rectal cancer. Six patients had a diverting ileostomy constructed as part of previous treatment for anastomotic complications before the salvage procedure. No anastomotic leaks were observed. All procedures but 1 were completed successfully. One patient who underwent DCA subsequently required an abdominoperineal resection and a permanent colostomy for postoperative extensive colonic ischemia. No 30-day mortality occurred.

Conclusion

Salvage Turnbull–Cutait DCA appears to be a safe procedure and could be offered to patients with complex anastomotic complications. This procedure could be added to the surgeon’s armamentarium as an alternative to the creation of a permanent stoma.     

Early lesions of follicular lymphoma: a genetic perspective

The pathogenesis of follicular lymphoma is a multi-hit process progressing over many years through the accumulation of numerous genetic alterations. Besides the hallmark t(14;18), it is still unclear which other oncogenic hits contribute to the early steps of transformation and in which precursor stages these occur. To address this issue, we performed high-resolution comparative genomic hybridization microarrays on laser-capture micro-dissected cases of follicular lymphoma in situ (n=4), partial involvement by follicular lymphoma (n=4), and duodenal follicular lymphoma (n=4), assumed to represent, potentially, the earliest stages in the evolution of follicular lymphoma. Cases of reactive follicular hyperplasia (n=2), uninvolved areas from follicular lymphoma in situ lymph nodes, follicular lymphoma grade 1–2 (n=5) and follicular lymphoma grade 3A (n=5) were used as controls. Surprisingly, alterations involving several relevant (onco)genes were found in all entities, but at significantly lower proportions than in overt follicular lymphoma. While the number of alterations clearly assigns all these entities as precursors, the pattern of partial involvement by follicular lymphoma alterations was quantitatively and qualitatively closer to that of follicular lymphoma, indicating significant selective pressure in line with its faster rate of progression. Among the most notable alterations, we observed and validated deletions of 1p36 and gains of the 7p and 12q chromosomes and related oncogenes, which include some of the most recurrent oncogenic alterations in overt follicular lymphoma (TNFRSF14, EZH2, MLL2). By further delineating distinctive and hierarchical molecular and genetic features of early follicular lymphoma entities, our analysis underlines the importance of applying appropriate criteria for the differential diagnosis. It also provides a first set of candidates likely to be involved in the cascade of hits that pave the path of the various progression phases to follicular lymphoma development.     

Clinical outcomes and risk factors for severe COVID-19 in patients with haematological disorders receiving chemo- or immunotherapy

Haematology patients receiving chemo- or immunotherapy are considered to be at greater risk of COVID-19-related morbidity and mortality. We aimed to identify risk factors for COVID-19 severity and assess outcomes in patients where COVID-19 complicated the treatment of their haematological disorder. A retrospective cohort study was conducted in 55 patients with haematological disorders and COVID-19, including 52 with malignancy, two with bone marrow failure and one immune-mediated thrombotic thrombocytopenic purpura (TTP). COVID-19 diagnosis coincided with a new diagnosis of a haematological malignancy in four patients. Among patients, 82% were on systemic anti-cancer therapy (SACT) at the time of COVID-19 diagnosis. Of hospitalised patients, 37% (19/51) died while all four outpatients recovered. Risk factors for severe disease or mortality were similar to those in other published cohorts. Raised C-reactive protein at diagnosis predicted an aggressive clinical course. The majority of patients recovered from COVID-19, despite receiving recent SACT. This suggests that SACT, where urgent, should be administered despite intercurrent COVID-19 infection, which should be managed according to standard pathways. Delay or modification of therapy should be considered on an individual basis. Long-term follow-up studies in larger patient cohorts are required to assess the efficacy of treatment strategies employed during the pandemic.     

Molecular profiling of Mycobacterium tuberculosis identifies tuberculosinyl nucleoside products of the virulence-associated enzyme Rv3378c

To identify lipids with roles in tuberculosis disease, we systematically compared the lipid content of virulent Mycobacterium tuberculosis with the attenuated vaccine strain Mycobacterium bovis bacillus Calmette–Guérin. Comparative lipidomics analysis identified more than 1,000 molecular differences, including a previously unknown, Mycobacterium tuberculosis-specific lipid that is composed of a diterpene unit linked to adenosine. We established the complete structure of the natural product as 1-tuberculosinyladenosine (1-TbAd) using mass spectrometry and NMR spectroscopy. A screen for 1-TbAd mutants, complementation studies, and gene transfer identified Rv3378c as necessary for 1-TbAd biosynthesis. Whereas Rv3378c was previously thought to function as a phosphatase, these studies establish its role as a tuberculosinyl transferase and suggest a revised biosynthetic pathway for the sequential action of Rv3377c-Rv3378c. In agreement with this model, recombinant Rv3378c protein produced 1-TbAd, and its crystal structure revealed a cis-prenyl transferase fold with hydrophobic residues for isoprenoid binding and a second binding pocket suitable for the nucleoside substrate. The dual-substrate pocket distinguishes Rv3378c from classical cis-prenyl transferases, providing a unique model for the prenylation of diverse metabolites. Terpene nucleosides are rare in nature, and 1-TbAd is known only in Mycobacterium tuberculosis. Thus, this intersection of nucleoside and terpene pathways likely arose late in the evolution of the Mycobacterium tuberculosis complex; 1-TbAd serves as an abundant chemical marker of Mycobacterium tuberculosis, and the extracellular export of this amphipathic molecule likely accounts for the known virulence-promoting effects of the Rv3378c enzyme.With a mortality rate exceeding 1.5 million deaths annually, Mycobacterium tuberculosis remains one of the world''s most important pathogens (1). M. tuberculosis succeeds as a pathogen because of productive infection of the endosomal network of phagocytes. Its residence within the phagosome protects it from immune responses during its decades long infection cycle. However, intracellular survival depends on active inhibition of pH-dependent killing mechanisms, which occurs for M. tuberculosis but not species with low disease-causing potential (2). Intracellular survival is also enhanced by an unusually hydrophobic and multilayered protective cell envelope. Despite study of this pathogen for more than a century, the spectrum of natural lipids within M. tuberculosis membranes is not yet fully defined. For example, the products of many genes annotated as lipid synthases remain unknown (3), and mass spectrometry detects hundreds of ions that do not correspond to known lipids in the MycoMass and LipidDB databases (4, 5).To broadly compare the lipid profiles of virulent and avirulent mycobacteria, we took advantage of a recently validated metabolomics platform (4). This high performance liquid chromatography–mass spectrometry (HPLC-MS) system uses methods of extraction, chromatography, and databases that are specialized for mycobacteria. After extraction of total bacterial lipids into organic solvents, HPLC-MS enables massively parallel detection of thousands of ions corresponding to diverse lipids that range from apolar polyketides to polar phosphoglycolipids. Software-based (XCMS) ion finding algorithms report reproducibly detected ions as molecular features. Each feature is a 3D data point with linked mass, retention time, and intensity values from one detected molecule or isotope. All features with equivalent mass and retention time from two bacterial lipid extracts are aligned, allowing pairwise comparisons of MS signal intensity to enumerate molecules that are overproduced in one strain with a false-positive rate below 1% (4).This comparative lipidomics system allowed an unbiased, organism-wide analysis of lipids from M. tuberculosis and the attenuated vaccine strain, Mycobacterium bovis Bacillus Calmette–Guérin (BCG). BCG was chosen because of its worldwide use as a vaccine and its genetic similarity to M. tuberculosis (6). We reasoned that any features that are specifically detected in M. tuberculosis might be clinically useful as markers to distinguish tuberculosis-causing bacteria from vaccines. Furthermore, given the differing potential for productive infection by the two strains, any M. tuberculosis-specific compounds would be candidate virulence factors. Comparative genomics of M. tuberculosis and BCG successfully identified “regions of deletion” (RD) that encode genes that were subsequently proven to promote productive M. tuberculosis infection (7), including the 6-kDa early secreted antigenic target (ESAT-6) secretion system-1 (ESX-1) (8, 9). We reasoned that a metabolite-based screen might identify new virulence factors because not all functions of RD genes are known. Also, biologically important metabolites could emerge from complex biosynthetic pathways that cannot be predicted from single-gene analysis.Comparison of M. tuberculosis and BCG lipid profiles revealed more than 1,000 differences, among which we identified a previously unknown M. tuberculosis-specific diterpene-linked adenosine and showed that it is produced by the enzyme Rv3378c. Previously, Rv3378c was thought to generate free tuberculosinol and isotuberculosinol (10–12). This discovery revises the enzymatic function of Rv3378c, which acts as a virulence factor to inhibit phagolysosome fusion (13). Whereas current models of prenyl transferase function emphasize iterative lengthening of prenyl pyrophosphates using one binding pocket, the crystal structure of Rv3378c identifies two pockets in the catalytic site, establishing a mechanism for heterologous prenyl transfer to nonprenyl metabolites.