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R J Walter

Publications and source records attributed to R J Walter.

At least 37 records · Page 2Linked to original sources
PubMed1994

Steroid sex hormones and macrophage function: modulation of reactive oxygen intermediates and nitrite release.

PROBLEM: In general, females have a more active immune response than do males. The effects of female sex hormones on lymphocytes have been studied extensively but their effects on macrophages are poorly understood. METHOD: In this study, peritoneal macrophages (M phi) obtained from male rats were treated in vitro with estradiol (E2), progesterone (P), testosterone (TS), or hydrocortisone (HC) and their effects on superoxide, hydrogen peroxide, and nitrite release determined. RESULTS: At concentrations between 10(-10) and 10(-9) M, female and male sex hormones had no significant effect on superoxide release but, at concentrations above or below that range, these hormones stimulated the release of these reactive oxygen intermediates (ROI). In contrast, M phi treated with HC generally exhibited either unaltered or reduced ROI release. CONCLUSIONS: These findings suggest that female sex hormones regulate ROI release by M phi in a manner not entirely shared by other steroid hormones. At most concentrations used, E2, P, TS, and HC significantly inhibited nitrite release by M phi. However, with 10(-10) M of E2 or 10(-9)M of P, nitrite release by M phi was not affected. In the presence of anti-TNF antibody, the amounts of superoxide and hydrogen peroxide release were moderately reduced but nitrite release was dramatically inhibited. The sensitivity of M phi to variations in the concentrations of female sex hormones may contribute to gender-related differences in the immune response.

Animals↗
PubMed1993

Modulation of the cytoskeleton and intracellular calcium in leukocytes exhibiting a cancer-associated chemotaxis defect.

Monocyte chemotaxis is severely depressed in patients with advanced tumors, but the cellular basis for this chemotactic defect is not known. Because the actomyosin cytoskeleton is thought to play a primary role in chemotaxis, we have employed flow cytometry to examine several aspects of the contractile machinery including myosin II, myosin light chain kinase (MLCK), actin, and cytoplasmic calcium in unstimulated and in formylpeptide-stimulated neutrophils and monocytes. Serum-pretreated polymorphonuclear leukocytes (PMNs) and monocytes from healthy blood donors or PMNs and monocytes isolated from tumor patients were studied. Leukocytes pretreated with serum from cancer patients exhibited decreased baseline myosin staining and a vastly different response to formylpeptide stimulation compared with leukocytes pretreated with normal human serum. In contrast, similar amounts of MLCK were observed in neutrophils and monocytes preincubated with normal or cancer serum with or without stimulation with formylpeptide. The fluorescent calcium indicator fluo-3 showed that resting and fMLP-stimulated levels of intracellular calcium were not significantly different in control and cancer serum-pretreated human leukocytes or in leukocytes isolated from tumor patients. Similarly, resting and fMLP-stimulated levels of F-actin in cancer patients' leukocytes as assessed by NBD-phallacidin staining did not differ significantly from those of normal leukocytes. Because the actomyosin cytoskeleton is intricately involved in leukocyte chemotaxis, alterations in the cytoskeleton may dramatically affect cell motility. The cytoskeletal alterations and changes in the response of leukocytes pretreated with cancer patients' serum to formylpeptide stimulation as described here may result in decreased chemotaxis by these cells.

Actins↗
PubMed1992

Newcastle disease virus selectively kills human tumor cells.

Newcastle disease virus (NDV), strain 73-T, has previously been shown to be cytolytic to mouse tumor cells. In this study, we have evaluated the ability of NDV to replicate in and kill human tumor cells in culture and in athymic mice. Plaque assays were used to determine the cytolytic activity of NDV on six human tumor cell lines, fibrosarcoma (HT1080), osteosarcoma (KHOS), cervical carcinoma (KB8-5-11), bladder carcinoma (HCV29T), neuroblastoma (IMR32), and Wilm's tumor (G104), and on nine different normal human fibroblast lines. NDV formed plaques on all tumor cells tested as well as on chick embryo cells (CEC), the native host for NDV. Plaques did not form on any of the normal fibroblast lines. To detect NDV replication, virus yield assays were performed which measured virus particles in infected cell culture supernatants. Virus yield increased 10,000-fold within 24 hr in tumor and CEC supernatants. Titers remained near zero in normal fibroblast supernatants. In vivo tumoricidal activity was evaluated in athymic nude Balb-c mice by subcutaneous injection of 9 x 10(6) tumor cells followed by intralesional injection of either live or heat-killed NDV (1.0 x 10(6) plaque forming units [PFU]), or medium. After live NDV treatment, tumor regression occurred in 10 out of 11 mice bearing KB8-5-11 tumors, 8 out of 8 with HT-1080 tumors, and 6 out of 7 with IMR-32 tumors. After treatment with heat-killed NDV no regression occurred (P less than 0.01, Fisher's exact test). Nontumor-bearing mice injected with 1.0 x 10(8) PFU of NDV remained healthy. These results indicate that NDV efficiently and selectively replicates in and kills tumor cells, but not normal cells, and that intralesional NDV causes complete tumor regression in athymic mice with a high therapeutic index.

Animals↗
PubMed1991

Hypophysectomy inhibits the synthesis of tumor necrosis factor alpha by rat macrophages: partial restoration by exogenous growth hormone or interferon gamma.

We recently demonstrated that GH and interferon-gamma (IFN gamma) act in a similar manner to prime macrophages in vitro and in vivo for enhanced superoxide anion release. In this report we investigated the physiological role of the pituitary gland and GH in in vivo priming of resident peritoneal macrophages for the synthesis of tumor necrosis factor-alpha (TNF alpha) in vitro. Compared to normal rats, hypophysectomized animals had an 83% reduction in macrophage production of TNF alpha after in vitro stimulation with lipopolysaccharide. Sham operation had no significant effect on the ability of macrophages to secrete TNF alpha in response to lipopolysaccharide. Both native pituitary-derived porcine GH (48 micrograms/rat.9 days) and native pituitary-derived rat GH (96 micrograms/rat.9 days) more than tripled the in vitro production of TNF alpha by macrophages from hypophysectomized rats (342 and 358 vs. 112 U/mg protein for placebo-treated rats, respectively). Each of these preparations of GH also increased growth more than 6-fold in hypophysectomized rats (32 and 30 g vs. 5 g in placebo controls). Heat inactivation of native pituitary-derived porcine GH significantly reduced its in vivo ability to augment both TNF alpha synthesis by macrophages and body growth. Recombinant rat IFN gamma (2000 U/rat.9 days) more than tripled the production of TNF alpha by macrophages from hypophysectomized rats (343 vs. 112 U/mg protein). In contrast to its in vivo effects, addition of GH in vitro to macrophages from hypophysectomized rats did not prime these cells for the synthesis of TNF alpha, indicating an indirect mechanism of action for GH. To further test the biological relevancy of GH with respect to synthesis of TNF alpha, hemorrhagic necrosis of TNF alpha-sensitive murine methyl-cholanthrene-induced tumors was assessed in pituitary-intact mice. Native porcine GH (133 micrograms/mouse.7 days) significantly augmented both the necrosis to tumor ratio and the hemorrhage to tumor ratio. These findings establish the physiological relevance of the pituitary gland and GH in the priming of macrophages for TNF alpha synthesis.

Animals↗
PubMed1990

Characterization of a chemotactic defect in patients with Kartagener syndrome.

Kartagener syndrome (KS) is an autosomally inherited recessive condition characterized by situs inversus, bronchiectasis, and chronic sinusitis. Ciliary dynein, the mechanochemical force generator in ciliary movement, is deficient in patients with KS. We examined blood samples from two patients and tissue biopsy specimens from five patients and found: (1) no significant defect in neutrophil or monocyte chemotaxis in response to formylpeptide chemoattractant; (2) no alterations in centriolar structure, but significantly more centriole-associated microtubules in KS neutrophils and monocytes than in control leukocytes; and (3) a marked reduction in KS fibroblast chemotaxis in response to fibronectin compared with control fibroblasts. The significance of these cellular defects in KS is described.

Adenosine Triphosphatases↗
PubMed1990

In vivo effects of recombinant interferon-gamma: augmentation of endotoxin-induced necrosis of tumors and priming of macrophages for tumor necrosis factor-alpha production.

Recombinant interferon-gamma (rIFN-gamma) is currently undergoing clinical trials in cancer patients. In this paper, we assessed the in vivo antitumor effects of this lymphokine in rodents. Recombinant murine IFN-gamma or control medium was injected intraperitoneally for 5 days into mice with subcutaneous Meth A tumors. An injection of a suboptimal dose of endotoxin (2 micrograms) on the fifth treatment day caused significant necrosis of tumors in the IFN-gamma-treated group while causing essentially no necrosis of tumors in the control group. Next, we examined macrophages isolated from rats treated for 9 days with either IFN-gamma or saline. Endotoxin stimulated release of significantly higher amounts of TNF-alpha from macrophages from the IFN-gamma-treated group compared to macrophages from the control group. A polyclonal antiserum against recombinant murine TNF-alpha abrogated all of the TNF cytotoxic activity from these rat macrophage supernatants, while control rabbit serum had no effect. These results provide strong evidence that rIFN-gamma can prime macrophages in vivo for TNF-alpha synthesis.

Animals↗
PubMed1989

Use of a laser-induced optical force trap to study chromosome movement on the mitotic spindle.

A laser-induced optical force trap was used to alter the movement of chromosomes in mitotic cells in vitro. The trap was produced by using a 1.06-microns neodymium YAG (yttrium/aluminum garnet) laser focused through a phase-contrast microscope. The trap was applied to one side of centrophilic chromosomes off the mitotic spindle and to late-moving chromosomes on the mitotic spindle. In both situations, chromosome movement was initiated in the direction opposite to that of the applied force. When the force was applied, chromosomes moved at velocities 10-20 times normal. These studies verify and extend the feasibility of using this new technique to study factors that influence organelle motility.

Animals↗
PubMed1989

Autoradiographic analysis of formylpeptide chemoattractant binding, uptake and intracellular processing by neutrophils.

Formylpeptide chemoattractant binds specifically to leukocyte cell surface receptors and during chemotactic activation, is internalized and processed by these cells. Using electron microscope autoradiographic techniques, we have examined the binding, uptake and disposition of fNle-Leu-Phe-Nle-[125I]Tyr-Lys by rabbit peritoneal neutrophils. Cells were incubated with ligand for 15 min at 4 degrees C, rinsed and then further incubated in buffer at 4 degrees C, 15 degrees C, 24 degrees C, or 37 degrees C for 0-40 min. For all cells incubated at 4 degrees C, grains were predominantly localized on the plasma membrane. Initially, this formylpeptide was seen in small clusters or microaggregates that were not associated with coated pits. Upon further incubation at 37 degrees C for 1 min, formylpeptide clustering on the plasma membrane increased and extensive internalization of formylpeptide was observed. Endocytosis of formylpeptide-receptor complexes clearly involved uncoated membrane pits and vesicles but did not appear to involve coated pits or coated vesicles. In the following 3 min, peptide proceeded in waves through compartments composed of small uncoated endocytic vesicles, then large vesicles, and then into dense granules. After 4 min at 37 degrees C, the most active phase of intracellular processing subsided. The percentage of grains in the cytoplasm, granule and small vesicle compartments very gradually increased during the remainder of the 40 min incubation. Formylpeptide neither bound at 4 degrees C nor accumulated at 37 degrees C on the cell surface in proximity to the underlying Golgi/centrosome region of the cell. At 24 degrees C, processing was slowed but formylpeptide-receptor complexes proceeded through a similar series of compartments. The t1/2 for formylpeptide uptake at 37 degrees C was 15-20 s, whereas at 24 degrees C the t 1/2 was approximately 10 min. No uptake was observed at 15 degrees C. The distinctive characteristics of formylpeptide binding and receptor-complex uptake seen here may be essential in initiating and maintaining continued chemotactic responsiveness.

Animals↗
PubMed1989

Viruses and annulate lamellae in Friend erythroleukemia cells.

Virus formation in a clone of murine undifferentiated Friend erythroleukemia cells was examined by electron microscopy. Budding C-type particles were present at the cell surface. The principal site of intracisternal particle production was in elements of rough-surfaced endoplasmic reticulum disposed about the periphery of stacks of annulate lamellae. Serial sections demonstrated that these virus-laden cisternae were in direct continuity with the annulate lamellae. In addition, intracisternal particles occurred in membranous honeycomb structures present in the cytoplasm of many cells. Viral elaboration also was associated with stacks of cisternae of endoplasmic reticulum that were devoid of ribosomes, but that were coated with an extensive and continuous layer of dense material. In some instances, the outer nuclear membrane was coated with the same dense substance. It appears that in Friend erythroleukemia cells, a very substantial portion of their cytomembranes is devoted to synthesis of intracisternal particles.

Animals↗
PubMed1988

Laser-transected microtubules exhibit individuality of regrowth, however most free new ends of the microtubules are stable.

To study the possible mechanism of microtubule turnover in interphase cells, we have used the 266-nm wavelength of a short-pulsed Nd/YAG laser to transect microtubules in situ in PtK2 cells at predefined regions. The regrowth and shrinkage of the transected microtubules have been examined by staining the treated cells with antitubulin mAb at various time points after laser irradiation. The results demonstrate that microtubules grow back into the transected zones individually; neither simultaneous growth nor shrinkage of all microtubules has been observed. The half-time of replacement of laser-dissociated microtubules is observed to be approximately 10 min. On the other hand, exposure of the core of the microtubule, which is expected to consist almost completely of GDP-tubulin, by transecting the internal regions of the microtubule does not render the remaining polymer catastrophically disassembled, and most transected microtubules with free minus ends do not quickly disappear. Taken together, these results suggest that most microtubules in cultured interphase cells exhibit some properties of dynamic instability (individual regrowth or shrinkage); however, other factors in addition to the hydrolysis of GTP-tubulin need to be involved in modulating the dynamics and the stability of these cytoplasmic microtubules.

Animals↗
PubMed1987

Direct visualization of formylpeptide receptor binding on rounded and polarized human neutrophils: cellular and receptor heterogeneity.

We have used light microscope autoradiography to visualize binding of the formylhexapeptide, N-formyl-norleucyl-leucyl-phenylalanyl-norleucyl-(125I)tyrosyl-lysine to rounded and spontaneously polarized human polymorphonuclear leukocytes. These cells possess receptors known to bind with high specificity and great avidity to the chemotactic formylpeptides. Cells adherent to glass slides were exposed to (125I)-hexapeptide at 4 degrees C, fixed, and autoradiographed. Hexapeptide binding was studied over the biologically active range of peptide concentrations varying from 0.63 nM to 10 nM and autoradiographic silver grains counted on 200 rounded or 50 polarized cells at each concentration. Examination of histograms plotted from these data revealed for rounded cells: two major peaks at each concentration indicating the existence of two neutrophil subpopulations, the predominant subpopulation binding one-half as much formylpeptide (peak I) as the other (peak II); progressively increasing proportions of cells in peak II as the free hexapeptide concentration increased. Accordingly, at 0.63 nM hexapeptide, peak II comprised only 8% of the total cell number, whereas at 10 nM, this peak represented 35% of the total cells. This suggested that different types of receptors may exist in the two cell subpopulations (high/low affinity or high/low negative cooperativity) and that these receptor types were expressed differentially on these subpopulations. Thus, cellular heterogeneity within the neutrophil population and receptor heterogeneity among hexapeptide receptors on an individual cell were both observed here. Each of these may significantly affect neutrophil functional responses to the chemotactic formylpeptides and may explain, at least in part, the curvilinearity in the Scatchard plots of formylpeptide receptor binding that has recently been reported. At higher concentrations of peptide (greater than or equal to 5 nM), spontaneously polarized PMN bound hexapeptide more or less uniformly over the entire cell surface. However, at lower concentrations, hexapeptide binding was markedly shifted toward the cell anterior. As a group, polarized PMN bound similar total quantities of hexapeptide, as did rounded PMN at each peptide concentration tested. Receptors displaying high- and low-affinity characteristics were, however, distributed asymmetrically over the cell surface, with the high-affinity type receptors predominantly on the anterior one-half of the cell. Such an asymmetric distribution may serve to initiate or perpetuate cell locomotion.

Humans↗
PubMed1987

Characterization of formylpeptide chemoattractant binding on neutrophils and monocytes from patients with head and neck cancer.

Monocytes from patients with head and neck tumors display a marked chemotactic deficiency in response to formylpeptide chemoattractants. Normal leukocytes possess receptors known to bind to chemotactic formylpeptides such as the hexapeptide N-formyl-nor-leucyl-leucyl-phenylalanyl-nor-leucyl-tyrosyl-lysine, with high specificity and great avidity. Light microscopic autoradiography was used to visualize the binding and subsequent cellular processing of the formylhexapeptide N-formyl-nor-leucyl-leucyl-phenylalanyl-nor-leucyl-[125I]tyrosyl-lysine to polymorphonuclear leukocytes (PMN) and monocytes from patients with epidermoid tumors of the head and neck. Surface-adherent leukocytes were exposed to 5 nM 125I-labeled hexapeptide for 15 minutes at 4 degrees C and were either fixed immediately or rinsed and then further incubated for 20 minutes at 37 degrees C prior to fixation. Cells underwent autoradiography, and silver grains associated with 400 cells were counted. There were no significant differences between the numbers of formylpeptide receptors on control and cancer patient PMN. However, there were significantly larger numbers of formylpeptide receptors (P less than .02) on cancer patient monocytes than on control monocytes. Cells that were allowed to process bound hexapeptide at 37 degrees C prior to fixation showed large decreases in the number of cell-associated silver grains. However, the amount of hexapeptide associated with PMN and monocytes treated in this way was the same for both control and tumor patients. In addition, histograms were plotted showing the amount of hexapeptide bound versus the frequency of cells at each level of binding. Examination of these histograms revealed that two major peaks or subpopulations occurred in each for neutrophils and monocytes from control patients. Histograms of grain counts for neutrophils and monocytes from tumor patients did not display two separate peaks of formylhexapeptide binding; instead, only one broad peak was evident. These observations indicated that: Tumor patient PMN expressed as many formylpeptide receptors as did control PMN, and tumor patient monocytes possessed significantly more receptors than did control monocytes. The disposition of formylpeptide initially bound to control and tumor patient leukocytes appeared to be essentially the same. Subpopulations existed in each control neutrophil and monocyte sample, one exhibiting approximately 30% more formylpeptide binding than the other. However, in the leukocytes obtained from tumor patients, the distinction between these two subpopulations had become obscured such that only one cell population was evident.

Carcinoma, Squamous Cell↗
PubMed1986

Clostridial collagenase. A chemoattractant for human neutrophils.

Leukocyte chemoattractants markedly alter the morphology and membrane functions of leukocytes. Bacterial collagenase causes a change in cell shape similar to that seen with the leukocyte chemoattractant, f-Met-Leu-Phe, and also promotes capping of concanavalin A. Human neutrophils in suspension or adherent to cover glasses were exposed to clostridial collagenase (10-250 units/ml) for up to 30 min at 37 degrees C and then fixed. Collagenase (125 units/ml) caused more than 85% of PMNs to assume an asymmetric or motile morphology even in the presence of 1% gelatin or 10 mg/ml bovine serum albumin. Trypsin alone (0.01-1%) did not induce a shape change. A similar morphology was seen in some untreated PMNs (less than 5% of all cells) and is characteristic of f-Met-Leu-Phe-treated cells (more than 90%). Collagenase inhibitors (i.e., reduced glutathione, cysteine, and acid-soluble collagen), however, prevented the shape change induced by collagenase but not by f-Met-Leu-Phe. At 4 degrees C, fluorescein-Con A (20 micrograms/ml) bound uniformly to both untreated and collagenase-treated cells. Upon further incubation at 37 degrees C, Con A was internalized over the entire cell periphery of the rounded, untreated cells but on collagenase-treated PMNs was rapidly gathered into a cap overlying the uropod or protuberant region of cytoplasm where it was subsequently internalized. Checkerboard Boyden chamber assays showed clostridial collagenase to be chemokinetic and chemotactic for human PMNs. In receptor binding experiments, the clostridial collagenase preparation competed poorly with [125I]formylhexapeptide for binding to PMN formylpeptide receptors (less than 15% reduction in binding at 200 units/ml collagenase). Thus, collagenase does not seem to interact strongly with the neutrophil formylpeptide receptor and may stimulate PMN motility by interacting at an altogether different site.

Cells, Cultured↗
PubMed1986

Increased viability and differentiation of normal and dystrophic striated muscle in vitro.

Primary cultures of muscle from normal (line 412) and dystrophic (line 413) chick embryos were exposed to corticosterone-21-acetate (C-21-A) or sodium ibuprofen (Motrin) for 28 d after myotube formation. Ibuprofen (0.5 to 500 micrograms/ml) or C-21-A (0.4 to 40 micrograms/ml)-treated cultures were fixed and assessed semiquantitatively using phase microscopy. On this basis, ibuprofen (50 micrograms/ml) and C-21-A (40 micrograms/ml) seemed to be effective in maintaining both normal and dystrophic muscle cultures. Using ibuprofen and C-21-A at these concentrations, experiments were repeated and analyzed quantitatively. Ibuprofen maintained culture viability (up to 68% more myotubes than untreated controls) but had no significant effect on the number of striated cells. C-21-A effectively maintained culture viability (up to 73% increase) and strongly promoted the formation of striated cells in these cultures (up to a sixfold increase). Both normal and dystrophic cultures were affected similarly by these agents, but the dystrophic cultures showed more consistent if not more extensive improvements in the parameters examined here. Thus, it seems that ibuprofen and C-21-A may affect both normal and dystrophic muscle directly to maintain survival and even promote differentiation.

Animals↗
PubMed1985

Brain dopamine and serotonin receptor sites revealed by digital subtraction autoradiography.

Autoradiography combined with image analysis permitted quantitative visualization of dopamine (D2) and serotonin (S2) binding sites in rat brain. Forebrain sections were incubated with tritiated spiroperidol alone or with tritiated spiroperidol plus unlabeled compounds that saturated the D2 or S2 sites. By subtracting the digitized image of an autoradiograph derived from the latter sections from that of the former, the D2 or S2 sites were specifically revealed. The resulting quantitative images demonstrate the differing anatomical distributions of these sites. The D2 site is largely restricted to the striatal complex (caudate-putamen, nucleus accumbens septi, and olfactory tubercle), whereas the S2 site is enriched in layer 5 of motor cortex, the perirhinal and cingulate cortices, and the claustrum.

Animals↗
PubMed1985

Defective monocyte chemotaxis in patients with epidermoid tumors of the head and neck.

The function of the immune surveillance system has been studied extensively in patients with neoplastic disease. Defective monocyte chemotaxis has been demonstrated in a variety of neoplastic conditions. We have assayed chemotaxis in monocytic and polymorphonuclear leukocytes taken from patients with epidermoid tumors of the head and neck. Multiwell chemotaxis chambers and the chemoattractant, formyl-methionyl-leucyl-phenylalanine, were used. Polymorphonuclear leukocytes responded normally, but monocytes from all tumor patients studied displayed marked reductions in random migration (86% inhibition) and in chemotaxis (77% inhibition) toward formylpeptide.

Adult↗
PubMed1985

Ability of specific monoclonal antibodies and conventional antisera conjugated to hematoporphyrin to label and kill selected cell lines subsequent to light activation.

A variety of cell lines have been prepared by fusion of the murine WEH1 3B cell line with peripheral blood leukocytes from a patient with chronic granulocytic leukemia. Fusion products were selected for their ability to produce a leukemia-associated antigen (CAMAL) previously described. One such line which originally produced CAMAL subsequently lost this ability and was used as a negative control. A number of antibodies were conjugated to hematoporphyrin (HP) and tested for their ability to bind to cell lines as detected by either fluorescence or by their ability to kill cells after light activation. The antibodies used were: rabbit anti-Hu (a conventional rabbit antiserum raised to membrane preparations from normal human peripheral blood leukocytes which served as a positive control); CAMAL-1 (a monoclonal gamma 1 antibody with specificity for the CAMAL antigen); and L1210 (an irrelevant monoclonal gamma 1 antibody). HP was conjugated to the antibodies by a carbodiimide procedure. When labeled cells were examined by fluorescence microscopy, it was apparent that both the rabbit antibody and CAMAL-1:HP showed positive labeling. The ability of the antibody:HP conjugates to kill labeled cells following light activation was tested. It was shown that rabbit anti-Hu:HP and CAMAL-1:HP conjugates were capable of killing significant numbers of cells when HP concentrations were as low as 1.2 ng/10(6) cells, whereas similarly treated cells exposed to either L1210:HP or HP alone did not exhibit significant killing until concentrations reached 240 and 120 ng/10(6) cells, respectively. Further experiments in which other cell lines were tested, all at HP concentrations of 12 ng/10(6) cells, demonstrated that those lines producing CAMAL were killed, whereas negative lines were not.

Animals↗